Influence of m6A methylation of influenza A virus RNA on the host innate immune response – METHAFLU

Today, there is a real revolution in therapeutic applications using or targeting nucleic acids (mRNA vaccines, antisense oligonucleotides (ASO) or siRNA). In addition, the role of RNA methylation and its diversity in human disease is emerging alongside that of DNA. The impact of DNA hypermethylation in certain cancers is well known, leading to the successful use of MTase inhibitors in leukaemia (AML). Deregulation of RNA methylation has been linked to the development of diseases such as cancer, and several pharmaceutical companies have focused their research on developing inhibitors of the METTL3/METTL14 complex, which catalyzes the N6 methylation of adenosine (m6A) of mRNA. More recently, it has been shown that RNA m6A modification plays a central role in the virus cycle. These methylations, introduced by the cellular methylase METTL3/14, which have various consequences on RNA structure, splicing, stability or translation, they enable some viruses to escape detection by the innate immune system, while they are detrimental for others. A preliminary study showed the importance of m6A modifications in the replication of influenza A virus (IAV), and proposed a global mapping of these modifications on IAV RNAs. However, this study failed to identify the pathways and mechanisms affected, notably due to the poor resolution of the mapping. So there are still numerous aspects that require further exploration.Our project proposes the parallel use of innovative experimental technologies and a statistical study of available sequences employing artificial intelligence to detect crucial viral RNA methylation sites for IAV replication and interspecies transmission. We will develop chemical tools that probe, inhibit or mimic these modifications in a context of viral infection. This will enable us to employ a chemobiological approach to identify the effects of m6A methylations on viral replication and production of infectious particles, the initiation of the innate immune response, and the metabolism of viral RNAs. Combining these results, we will experimentally determine the influence of methylation on the triggering of the innate immune response in various state-of-the-art cell systems, notably using both multicellular models of human lung epithelium in vitro and ex vivo models of murine and human lung tissue culture. We will then identify the cellular players involved by exploring the structural dynamics of RNA induced by methylation, by identifying the proteins bound to methylated RNA and, finally, by reverse genetics and designing mutant viruses. From a chemical perspective, this proposal aims to synthesize inhibitors of viral methyltransferases using novel and original scaffolds, designed through molecular modeling and validated through in vitro and in cellulo biological assays. This approach is expected to yield molecules with high selectivity and efficacy.
Our multidisciplinary study will shed unique light on host-pathogen relationships, and we hope to define new therapeutic targets whose scope could extend beyond IAV viruses.