The epitranscriptome in Drosophila melanogaster

• Applicant: Professor Dr. Mark Helm
• Subject Area: General Genetics and Functional Genome Biology ; Biochemistry ; Developmental Biology
• Term: from 2017 to 2021
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 393465853

The field of RNA modifications, now termed Epitranscriptomics, has made major strides to the central stage of gene expression control in the recent years. Many modified nucleotides were first discovered in transfer RNAs and ribosomal RNAs in all studied organisms. Few of the same nucleotide modifications were later identified in messenger RNAs (mRNA) of eukaryotes, where they were recently revealed as an entire new layer of regulation of gene expression. The most prevalent and best-studied modification on mRNA is N6-methyladenosine (m6A). This modification was shown to regulate several physiological processes in various organisms via posttranscriptional control of gene expression. We recently characterized the m6A pathway in Drosophila melanogaster by identifying its main components and demonstrating its critical role during neurogenesis and sex determination. While this work substantially advanced our understanding on m6A biogenesis and function, it also opened new questions while revealing several limitations. For instance it remains unclear how, and through which components the methyltransferase complex (m6A writer) recognizes the putative m6A methylation sites and why only few of these sites are methylated. So far, five different factors were shown to belong to the methyltransferase complex; however their exact function is still unclear and currently none of them convincingly explains how the specificity of the complex is achieved. In addition, our m6A profiling via antibodies highlighted the need for the development of more robust and less biased approaches to precisely map m6A on mRNA with high confidence. In this collaborative proposal we want i) to obtain insights into the composition and mode of action of the m6A methyltransferase complex, ii) to develop new single resolution mapping techniques for m6A residues, and lastly iii) to address whether other RNA modifications are present on mRNA in Drosophila and how they might be regulated. Our preliminary data revealed that additional factors likely belong to the methyltransferase complex, in particular the not yet characterized protein CG7358, and we propose to study its function within the complex. Furthermore, we have generated promising results in regards to the identification of m6A sites using an approach based on small molecule chemistry. This approach must be developed for the transcriptome wide level. Lastly our unpublished work suggests that 2-3 additional modifications could be present on mRNA in Drosophila. We plan to further characterize these modifications as well as to identify their regulatory players. The suggested cooperative study has the potential to greatly impact our understanding of the nascent field of mRNA modifications and to provide important insights into what is likely to be a sophisticated regulatory network.

• DFG Programme: Research Grants