The function of the RNA methylome in animals

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RNA modifications control diverse biological processes such as embryonic development, sex determination, nutrient sensing and many others. There are currently more than 150 distinct modifications on RNA. The diversity of RNA modifications alongside the proteins required for their synthesis, removal and recognition poses a significant challenge to understanding their biological role. This project aims to uncover (1) the function of RNA methylations in animal development and (2) how RNA methylations are regulated. Many types of RNA contain a diverse number of RNA modifications. Ribosomal and transfer RNAs are heavily modified. Discoveries of internal RNA modifications on messenger RNAs have further expanded the role RNA modifications play in gene expression regulation. Furthermore, several other non-coding RNAs are modified, such as spliceosomal snRNAs and many small regulatory RNAs. We use genetic and biochemical tools to understand how RNA modifications function during germline development and how diet affects RNA modification levels. We use state-of-the-art technologies, including Nanopore Sequencing and high-resolution mass spectrometry. Regulation of gene expression is crucial in the germline. Germ cells give rise to offspring, and gene expression changes in germ cells can directly affect embryonic development. In many organisms, including nematodes and mammals, gene expression changes in the germ cells can have multi-generational consequences. Our research focuses on how RNA modifications and RNA modifying enzymes regulate gene expression in animal germ cells using the nematode Caenorhabditis elegans as a discovery organism. During C. elegans germline development, germ cells proliferate from mitosis to meiosis and differentiate into oocytes or sperm. Therefore, we can use the C. elegans germline to investigate cell proliferation and differentiation at the same time. We will first characterise the role of RNA methyltransferases in mRNA maturation through the regulation of RNA splicing. mRNA maturation is a complex process that involves cap methylation, splicing of introns and poly-adenylation. Efficient RNA splicing requires orchestrating a large protein complex that contains many non-coding RNAs. In addition to removing introns, nematodes and many other organisms that include many parasitic species use trans-splicing during mRNA maturation. Trans-splicing replaces the 5' end of mRNAs with a non-coding SL-RNA. We will use direct RNA sequencing and biochemical approaches to understand how snRNA modifications regulate splicing processes in germ cells.Next, we will investigate how dietary vitamin B12 can regulate RNA modifications. Vitamin B12 is essential for the methionine cycle, which controls S-adenosylmethionine levels. Bacteria and archaea can synthesise vitamin B12, whereas animals must acquire vitamin B12 through their diet. Using our "vegan diet" system and isotope labelling, we will trace how RNA methylation levels change in the presence and absence of vitamin B12. We will use nanopore RNA sequencing to determine isoform level changes in gene expression in the presence and absence of vitamin B12. We will further expand our analysis to bacteria found in the natural diet of C. elegans. We will determine how C. elegans could sense vitamin B12 at the gene expression level. Our research is crucial because it will advance our understanding of how RNA modifications function during development and enable future research in this area by generating tools and resources. Our research addresses a much-overlooked topic connecting diet to chemical changes on RNA.