Essential roles of RNA cap-proximal ribose methylation in mammalian embryonic development and fertility

Eukaryotic RNA pol II transcripts are capped at the 5′ end by the methylated guanosine (m7G) moiety. In higher eukaryotes, CMTR1 and CMTR2 catalyse cap-proximal ribose methylations on the first (cap1) and second (cap2) nucleotides, respectively. These modifications mark RNAs as ‘self’, blocking the activation of the innate immune response pathway. Here we show that loss of mouse Cmtr1 or Cmtr2 leads to embryonic lethality, with non-overlapping sets of transcripts being regulated, without activation of the interferon pathway. In contrast, Cmtr1 mutant adult mouse livers exhibit a chronic activation of the interferon pathway, with multiple interferon-stimulated genes (ISGs) being expressed. Conditional deletion of Cmtr1 in the male germline leads to infertility, while global translation is unaffected in the Cmtr1 mutant mouse liver and human cells. Thus, mammalian cap1 and cap2 modifications have an essential role in embryonic gene regulation that is independent of their established role in evading the innate immune system. RNA sequencing was applied to study the gene expression differences between Cmtr1+/+ and Cmtr1-/- mouse embryos (E6.5 and E7.5), Cmtr1+/loxP; AlbCreERT2+/- (control) and Cmtr1-/loxP; AlbCreERT2+/- (Cmtr1 cKO) mouse livers, CMTR1+/+ and CMTR1-/- HAP1 cells, Cmtr2+/+ and Cmtr2-/- mouse embryos (E6.5 and E7.5). Ribosome profiling was used to assess translation efficiency differences between Cmtr1+/loxP; AlbCreERT2+/- (control) and Cmtr1-/loxP; AlbCreERT2+/- (Cmtr1 cKO) mouse livers, CMTR1+/+ and CMTR1-/- HAP1 cells.