Exploring antiviral RNA interference in mammals__Renewal
Lead Research Organisation:
Queen Mary University of London
Department Name: Blizard Institute of Cell and Molecular
Abstract
Viruses constitute a constant threat for humans as illustrated by the recent pandemic of COVID-19 caused by severe acute respiratory syndrome coronavirus 2 (SARS CoV 2). There is therefore an urgent need to understand the defence mechanisms that mammals have at their disposal to combat virus infections so that we can develop new antiviral therapies.
In plants, insects and worms, one major defence mechanism is called antiviral RNA interference (RNAi). Viruses can only reproduce after infecting a living cell. Once inside the cell, viruses hijack the cell's machinery to make new copies of their viral genome, which in many cases, are made of ribonucleic acid (RNA). The antiviral RNAi system uses a succession of molecular scissors to destroy viruses within the cells. First a protein from the cell, called Dicer, chops the viral genome when it is being copied into small inhibitory fragments. These small molecules of RNA stick to all the copies of the viral genome and act as tell-tale signs for another protein called Argonaute 2 (Ago2) to cleave these viral RNA copies. In mammals, cells are equipped with another defence mechanism called the interferon (IFN) system, which was long thought to have completely replaced the more ancestral RNAi-based system. However, recent data generated by myself and other labs have demonstrated that this antiviral RNAi is also active in mammals. I further uncovered that the IFN system acts as a brake on antiviral RNAi in certain cell types. These studies were carried out using in vitro cell culture and it is now important to test how much RNAi helps to protect living animals from infections.
Firstly, I will address how important this ancestral defence mechanism is by testing how much adult animals cope with infections if their molecular scissors, Ago2, is rendered defective. Secondly, I will assess the full antiviral potential of this RNAi-based defence by releasing the brake imposed by the IFN system and thereby unleashing its activity. For this, we will test how efficient RNAi is at protecting animals from virus infections in mice that lack the IFN system. Thirdly, I will test whether Ago2 cuts not only RNA molecules coming from viruses but also RNA molecules produced by the cells. RNAi activity is particularly high in stem cells, which are cells that are not yet completely specialised for a specific function and therefore can give rise to a number of different cell types. These stem cells are found either in the early embryo (embryonic stem cells, ESCs) or in various tissues of the body (tissue stem cells). Previous work in ESCs as well as in immature eggs (oocytes), both known to have potent RNAi activity, hints toward a possible mechanism by which Ago2 regulates expression of some genes by cleaving so-called messenger RNAs that are essential to relay the genetic information from genes to proteins. Some of these genes are transposons, which are ancient viruses that have invaded mammalian genomes over the course of evolution. These transposons are tightly regulated because they can jump from one part of the genome to another, which can have dramatic consequences if they land in and disrupt cellular genes. We will therefore test the exciting possibility that Ago2 acts as a molecular scissors to regulate gene expression and control transposons in cells endowed with high RNAi activity, i.e. in stem cells. We will examine ESCs and tissue stem cells from animals in which Ago2 scissors activity can be switched off and test the impact on expression of genes and transposons.
This research programme tackles fundamental and important questions to elucidate the functional role of this ancestral mode of defence in mammalian immunology. Knowledge generated will improve our understanding and ability to boost natural antiviral mechanisms. Moreover, the results will also be relevant to deciphering the fundamental biology of stem cells.
In plants, insects and worms, one major defence mechanism is called antiviral RNA interference (RNAi). Viruses can only reproduce after infecting a living cell. Once inside the cell, viruses hijack the cell's machinery to make new copies of their viral genome, which in many cases, are made of ribonucleic acid (RNA). The antiviral RNAi system uses a succession of molecular scissors to destroy viruses within the cells. First a protein from the cell, called Dicer, chops the viral genome when it is being copied into small inhibitory fragments. These small molecules of RNA stick to all the copies of the viral genome and act as tell-tale signs for another protein called Argonaute 2 (Ago2) to cleave these viral RNA copies. In mammals, cells are equipped with another defence mechanism called the interferon (IFN) system, which was long thought to have completely replaced the more ancestral RNAi-based system. However, recent data generated by myself and other labs have demonstrated that this antiviral RNAi is also active in mammals. I further uncovered that the IFN system acts as a brake on antiviral RNAi in certain cell types. These studies were carried out using in vitro cell culture and it is now important to test how much RNAi helps to protect living animals from infections.
Firstly, I will address how important this ancestral defence mechanism is by testing how much adult animals cope with infections if their molecular scissors, Ago2, is rendered defective. Secondly, I will assess the full antiviral potential of this RNAi-based defence by releasing the brake imposed by the IFN system and thereby unleashing its activity. For this, we will test how efficient RNAi is at protecting animals from virus infections in mice that lack the IFN system. Thirdly, I will test whether Ago2 cuts not only RNA molecules coming from viruses but also RNA molecules produced by the cells. RNAi activity is particularly high in stem cells, which are cells that are not yet completely specialised for a specific function and therefore can give rise to a number of different cell types. These stem cells are found either in the early embryo (embryonic stem cells, ESCs) or in various tissues of the body (tissue stem cells). Previous work in ESCs as well as in immature eggs (oocytes), both known to have potent RNAi activity, hints toward a possible mechanism by which Ago2 regulates expression of some genes by cleaving so-called messenger RNAs that are essential to relay the genetic information from genes to proteins. Some of these genes are transposons, which are ancient viruses that have invaded mammalian genomes over the course of evolution. These transposons are tightly regulated because they can jump from one part of the genome to another, which can have dramatic consequences if they land in and disrupt cellular genes. We will therefore test the exciting possibility that Ago2 acts as a molecular scissors to regulate gene expression and control transposons in cells endowed with high RNAi activity, i.e. in stem cells. We will examine ESCs and tissue stem cells from animals in which Ago2 scissors activity can be switched off and test the impact on expression of genes and transposons.
This research programme tackles fundamental and important questions to elucidate the functional role of this ancestral mode of defence in mammalian immunology. Knowledge generated will improve our understanding and ability to boost natural antiviral mechanisms. Moreover, the results will also be relevant to deciphering the fundamental biology of stem cells.
