Uncovering the role of the iRhom2-ADAM17 interaction in inflammatory signalling

Project Information

iRhomADAM

Grant agreement ID: 840146

DOI

10.3030/840146

Project closed

EC signature date 10 April 2019

Start date 1 September 2020

End date 1 April 2023

Funded under

• EXCELLENT SCIENCE - Marie Skłodowska-Curie Actions

Total cost

€ 212 933,76

EU contribution

€ 212 933,76

Coordinated by

THE CHANCELLOR, MASTERS AND SCHOLARS OF THE UNIVERSITY OF OXFORD
United Kingdom

Periodic Reporting for period 1 - iRhomADAM (Uncovering the role of the iRhom2-ADAM17 interaction in inflammatory signalling)

Reporting period: 2020-09-01 to 2022-08-31

Summary of the context and overall objectives of the project

Most of the cells in our body have the ability to detect damages and infections, and recruit the immune system to the "crime scene" to clear out the cause of harm. The immune system will then execute a number of action plans, the first one is inflammation. In normal, healthy situations, the cells only send signals to recruit the immune system when they are actually harmed, and they cease their signal once the damage was cleared. This way, inflammation only takes place when it is needed, and it remains restricted to the site and to the duration of the injury. However, in some situation, the regulation of inflammation is compromised, and immune cells will be active even when they are not actually needed. This can cause serious damage to healthy tissues. Therefore, it is important to understand the programs that cells use to recruit the immune system, as well as the programs the immune system uses to receive these signals. Once we decipher the details of these programs, i.e. which are the participating proteins, what activates each protein, what is the outcome that each protein activation has, etc; we can design treatments for conditions were the regulation of inflammation is lost.
This project aims to shed light on some of the action plans that cells use once they detect that they were infected with a virus. More specifically, in this project I am studying the role of a protein called iRhom2 in the cells' response to viruses. iRhom2 is an elusive protein that normally resides in membranes that encapsulate our cells or in membranes that are found inside the cells. Not much is known about its function, but previous work from our group and from other groups shows that iRhom2 is involved in cancer and in inflammation. My aim was to expand the knowledge on iRhom2 to the distinct immune action plan called "the antiviral response". So far my work is showing that while iRhom2 is important in inflammation, when it comes to antiviral activity, iRhom2 actually tunes that action plan down. I am currently still working on understanding exactly what iRhom2 does in this cellular program, with a special emphasis on what are the features that allow it to promote inflammation while suppressing antiviral activity. If iRhom2 is chosen as a drug target for chronic inflammation (inflammation that is not restricted in time and in place), it is essential to understand its full role in immunity, in order to make sure that the future drug has no serious side effects.

Work performed from the beginning of the project to the end of the period covered by the report and main results achieved so far

The project started with an unbiased approach of simulating viral infection in cultured mammalian cells. Since I wanted to understand the role of the protein iRhom2, I tested two lines of mouse cell lines - cells that have iRhom2 and cells that do not have iRhom2. Both of these lines were generated in our lab a few years ago and both were previously reported in the literature. In these experiments, cells that did not have iRhom2 had a much higher immune response to the treatment than cells that did have iRhom2, and it became clear that under these conditions, iRhom2 is restricting the immune response to viruses. These results were surprising, as they were somewhat contradicting previously published results about the role of iRhom2 in this type of immune response. Therefore I wanted to confirm that my results are robust and reproducible. For that I used a number of other cell lines, both from mouse and from human. All of which showed the same phenomenon. Once this was established, I was ready to move on and look at two parallel questions: 1. What is iRhom2 doing in the cell to promote this restricting activity? 2. What is the biological context by which iRhom2 is restricting antiviral activity while being an important regulator of inflammation, an essential immune response to many kinds of infections and other injuries. The latter is important both for our understanding of how different immune pathways regulate one another, and for for potential future therapeutics, where it would be essential to consider the full activity of iRhom2 in the immune system.
My approach to understanding how iRhom2 is operating in the cells was to use existing knowledge on the cell's response to viral infection to design experiments, without preexisting assumptions on what iRhom2 might be doing in this context. My results demonstrate that iRhom2 is acting before the massive production of the key protein interferon-beta, suggesting that iRhom2 could be involved in regulating its production. Additionally, I also saw that while the overall outcomes of inflammation and of antiviral activity happen simultaneously, iRhom2 is regulating each one of them in a different mechanism of action. I am currently working on pinpointing the exact role of iRhom2 in antiviral immunity. Specifically, what other proteins it interacts with and under what conditions.
In order to understand the wider immune context of iRhom2, I used a type of immune cells called macrophages, which were generated from mouse bone marrow by my colleagues in the lab. These cells were treated with the same treatment that simulates viral infection, and then their total RNA was isolated and sequenced, in order to characterise the differences between cells with iRhom2 and cells without it. So far, these cells exhibit significant changes in their overall immune activity, as well as their energy production. These results need to be validated in additional cell lines, and then combined with the mechanistic knowledge, to shed light about the global role of iRhom2 in this pathway.

Progress beyond the state of the art and expected potential impact (including the socio-economic impact and the wider societal implications of the project so far)

There are two directions that I work on in parallel. The first one is to define the molecular mechanism that governs iRhom2's activity in antiviral immunity. i.e what exactly does it do in the cell: what other proteins it interacts with? What cellular stimulus it responds to? etc. The second direction is to learn the wider impact of iRhom2 activation, in order to understand what is its full role in immunity. At this point we know that it is important for inflammation, while simultaneously restricting antiviral activity. It is well established that the full immune response is a multi-layered, intricate matrix, when different branches are activated by different cells, at different time points, in response to different stimuli. The overall outcome is determined by the sum of all of the activities. So far my results suggest that iRhom2 could be acting at one of the intersections of the multi-layered regulatory network of branches. After completing these two parts I will write a manuscript for a peer review, open access journal.
iRhom2 was suggested as a drug target for conditions of chronic inflammation. Understanding the full role of iRhom2 in the immune response will shed light on its true potential as a drug target, since it will provide essential information in order to predict side effects of that treatment. Having this knowledge will help us make educated decisions on how viable it actually is as a drug target, and secondly, we may be able to develop very specific treatments that will affect some functions and not others. This knowledge could save time and money on trials that are bound to fail due to significant side effects. This is particularly important when considering steps of pre-clinical experiments on animals: With better knowledge, some these experiments may turn out to be unnecessary and could be spared.

Suggested roles for iRhom2 activity in antiviral immunity