DOA/CLK2 phosphorylates Fl(2)d/WTAP to enhance m6A mRNA methyltransferase complex activity

Abstract

N6-methyladenosine (m6A) is the most abundant internal modification in messenger RNA (mRNA) providing an essential layer to the control of gene expression important to many biological processes. A megadalton writer complex deposits m6A methyl marks that are then read by YTH domain containing proteins. Deposition of m6A is thought to be regulated by cellular signalling but how remains uncertain. We identified the kinase DOA, a highly conserved homologue of human CLK2 by using the essential role of m6A in the Drosophila Sxl auto-regulation for sex determination and dosage compensation. We show that DOA kinase is required for m6A deposition and Sxl alternative splicing. Overexpression of DOA can compensate writer complex insufficiency and rescue lethality of m6A in the context of Sxl mis- splicing, suggesting a key role in regulating activity of the m6A complex. Through genetic interaction experiments, we identify a phospho-site at the end of a conserved helical structure in Fl(2)d as DOA target. CLK2 phosphorylates the same helix in WTAP, the human homologue of Fl(2)d. CLK2 expands regulatory capacity through additional phospho-sites and changing of the Drosophila site to a phosphomimetic-like glutamine, overall important for m6A directed regulation of dosage compensation genes in human cells.