Identification of N6-methyladenosine (m6A) modified RNA in ILC2 of WT and Mettl3 Klrg1Cre mice

Innate lymphoid cells (ILCs) are able to directly respond to alarmin signals and produce an array of effector molecules for immune protection and tissue homeostasis. However, how posttranscriptional machinery in ILCs execute extracellular stimuli towards robust gene expression is yet to understand. Here, we reported a cell type-specific role of N6-methyladenosine (m6A) RNA methylation in ILCs. Inducible deletion of m6A methyltransferase METTL3 had little impact on ILC maintenance in the steady state or cytokine-induced ILC1 or ILC3 activation, but dramatically diminished IL-25-triggered ILC2 response. Specific deletion of Mettl3 in ILC2 significantly attenuated cell expansion, cytokine production, inter-organ migration, and anti-helminth immunity. To investigate the molecular mechanism by which m6A modification regulates ILC2 response, we subjected ILC2 of WT and Mettl3 Klrg1Cre mice to a m6A-tagged mRNA immunoprecipitation sequencing (meRIP-seq) to identify the m6A modified mRNA. Overall design: We sorted ILC2 from small intestine of WT and Melttl3 Klrg1Cre mice and isolated the total RNA from the ILC2. Then we performed methylated RNA immunoprecipitation (meRIP) using anti-m6A antibody to obtain the pellet fractions which are m6A modified RNA fractions. We next constructed the libraries for pellet fractions and performed the sequencing.