A neural mA/Ythdf pathway is required for learning and memory in Drosophila.

Epitranscriptomic modifications can impact behavior. Here, we used Drosophila melanogaster to study N-methyladenosine (mA), the most abundant modification of mRNA. Proteomic and functional analyses confirm its nuclear (Ythdc1) and cytoplasmic (Ythdf) YTH domain proteins as major mA binders. Assays of short term memory in mA mutants reveal neural-autonomous requirements of mA writers working via Ythdf, but not Ythdc1. Furthermore, mA/Ythdf operate specifically via the mushroom body, the center for associative learning. We map mA from wild-type and Mettl3 mutant heads, allowing robust discrimination of Mettl3-dependent mA sites that are highly enriched in 5' UTRs. Genomic analyses indicate that Drosophila mA is preferentially deposited on genes with low translational efficiency and that mA does not affect RNA stability. Nevertheless, functional tests indicate a role for mA/Ythdf in translational activation. Altogether, our molecular genetic analyses and tissue-specific mA maps reveal selective behavioral and regulatory defects for the Drosophila Mettl3/Ythdf pathway.