Single-cell gene expression profiling revealed mechanisms with potential roles in regulating viral replication and vascular function. The findings provide new insights into host-virus interactions and establish single-cell sequencing as a valuable tool for fish research.
The endothelium (the inner cellular layer of blood vessels) is a central player in viral disease. This has been best demonstrated in avian influenza, where the ability to infect endothelial cells is closely linked to the virus' potential to cause disease. Many other viruses with high relevance to human and animal health (like henipaviruses, hantavirus, and bluetongue virus) infect and replicate in endothelial cells. However, most studies of the interplay between virus and endothelial cells have been conducted in two-dimensional cell cultures under laboratory conditions. Endothelial cells are strongly influenced by their microenvironment, emphasizing the importance of studies that highlight events during infection of live hosts. In addition to understanding factors that influence viral replication in endothelium, it is important to understand how virus infection compromises vascular function, leading to disturbed coagulation, loss of vascular integrity, and/or uncontrolled inflammation.
In this project we will have used infectious salmon anaemia virus (ISAV) as a model to understand the interplay between virus and endothelial cells during infection. We have sampled infected fish and profiled gene expression in thousands of individual cells, enabling identification of cell type-specific response patterns with focus on endothelial cells. Our aim has been to identify factors that promote or limit viral replication, detection by the immune system, and/or influence vascular functions. We have examined different stages of infection and compared the response to viruses that cause low and high mortality.
The project has confirmed that virus production in infected fish primarily occurs in the vascular endothelium and identified two possible mechanisms that limit viral replication in endothelial cells.
First, the virus destroys its own receptor when it binds to the cell, preventing new viruses from attaching. The loss of the receptor affects both endothelial cells and red blood cells and correlates with an increase in virus levels in the blood.
Second, we observe that endothelial cells exhibit the most significant changes in gene expression during infection. We have identified two response patterns in these cells. The first corresponds to high expression of viral RNA and represents the response in cells that permit viral replication. The second response pattern characterizes a group of cells that do not express viral RNA, suggesting that they restrict viral replication. This endothelial subpopulation shows high expression of several genes that have been shown to protect other cell types from infection.
We have also mapped the different cell types present in the head kidney, including subgroups of endothelial cells and immune cells, and how gene expression in each cell type changes during ISA virus infection. While this project has focused on the endothelial response, the data will be analyzed further in an EU-funded project to understand how immune cells contribute to the fish’s defense against ISAV.
In summary, the project has provided new insights into the interaction between the virus and endothelial cells in general, as well as important information about the response of farmed Atlantic salmon to viral infection. Additionally, the project has enabled the establishment of single-cell sequencing at the Norwegian Veterinary Institute, providing a powerful tool to investigate complex biological questions in fish and other animals, where cell-type-specific antibodies are often lacking.
Project manager
Partners
Wellcome Sanger Institute, UK
