N6 -methyladenosine (m6 A) is a prevalent epitranscriptomic mark in eukaryotic RNA, with crucial roles for mammalian and ecdysozoan development. Indeed, m6 A-RNA and the related protein machinery are important for splicing, translation, maternal-to-zygotic transition and cell differentiation. However, to date, the presence of an m6 A-RNA pathway remains unknown in more distant animals, questioning the evolution and significance of the epitranscriptomic regulation. Therefore, we investigated the m6 A-RNA pathway in the oyster Crassostrea gigas, a lophotrochozoan model whose development was demonstrated under strong epigenetic influence. Using mass spectrometry and dot blot assays, we demonstrated that m6 A-RNA is actually present in the oyster and displays variations throughout early oyster development, with the lowest levels at the end of cleavage. In parallel, by in silico analyses, we were able to characterize at the molecular level a complete and conserved putative m6 A machinery. The expression levels of the identified putative m6 A writers, erasers and readers were strongly regulated across oyster development. Finally, RNA pull-down coupled to LC-MS/MS allowed us to prove the actual presence of readers able to bind m6 A-RNA and exhibiting specific developmental patterns. Altogether, our results demonstrate the conservation of a complete m6 A-RNA pathway in the oyster and strongly suggest its implication in early developmental processes including MZT. This first demonstration and characterization of an epitranscriptomic regulation in a lophotrochozoan model, potentially involved in the embryogenesis, bring new insights into our understanding of developmental epigenetic processes and their evolution.
Keywords: RNA; development; epitranscriptomics; methylation; oyster.
© 2020 Federation of European Biochemical Societies.