A Dicer-independent miRNA biogenesis pathway that requires Ago catalysis Sihem Cheloufi, Camila O. Dos Santos, Mark M. W. Chong, and Gregory J. Hannon identified a Dicer-independent miRNA biogenesis pathway that requires Ago catalysis. In mice, Ago2 is uniquely required for viability and retains catalytic competence. They engineered a mouse with catalytically inactive Ago2 alleles, resulting in early embryonic lethality and loss of miR-451, a small RNA important for erythropoiesis. The pre-miRNA is loaded into Ago and cleaved by the Ago catalytic centre to generate an intermediate 3' end, which is then further trimmed. This pathway is conserved and important for vertebrate development. Argonaute proteins are key effectors of small RNA-mediated regulatory pathways. The PAZ and Mid domains help anchor the small RNA guide, while the Piwi domain contains an RNase H motif. Ago proteins are divided into three clades, with Ago2 being the only one retaining catalytic competence in mammals. Ago2 is essential for extra-embryonic development, as shown by the lethality of Ago2-null embryos. However, catalytic activity is required for postnatal development, as Ago2-null mice become pale and die shortly after birth. The study identified that miR-451 maturation requires Ago2 catalysis, as it is not processed by Dicer. miR-451 is essential for erythropoiesis, and its loss leads to anaemia. The study also showed that miR-451 biogenesis occurs through a non-canonical pathway, with Drosha cleavage followed by direct loading into Ago2. This pathway is conserved across vertebrates and may contribute to the evolutionary pressure to maintain a catalytic Argonaute protein. The findings suggest that Ago2 catalysis is essential for miR-451 maturation and that this pathway is important for vertebrate development.A Dicer-independent miRNA biogenesis pathway that requires Ago catalysis Sihem Cheloufi, Camila O. Dos Santos, Mark M. W. Chong, and Gregory J. Hannon identified a Dicer-independent miRNA biogenesis pathway that requires Ago catalysis. In mice, Ago2 is uniquely required for viability and retains catalytic competence. They engineered a mouse with catalytically inactive Ago2 alleles, resulting in early embryonic lethality and loss of miR-451, a small RNA important for erythropoiesis. The pre-miRNA is loaded into Ago and cleaved by the Ago catalytic centre to generate an intermediate 3' end, which is then further trimmed. This pathway is conserved and important for vertebrate development. Argonaute proteins are key effectors of small RNA-mediated regulatory pathways. The PAZ and Mid domains help anchor the small RNA guide, while the Piwi domain contains an RNase H motif. Ago proteins are divided into three clades, with Ago2 being the only one retaining catalytic competence in mammals. Ago2 is essential for extra-embryonic development, as shown by the lethality of Ago2-null embryos. However, catalytic activity is required for postnatal development, as Ago2-null mice become pale and die shortly after birth. The study identified that miR-451 maturation requires Ago2 catalysis, as it is not processed by Dicer. miR-451 is essential for erythropoiesis, and its loss leads to anaemia. The study also showed that miR-451 biogenesis occurs through a non-canonical pathway, with Drosha cleavage followed by direct loading into Ago2. This pathway is conserved across vertebrates and may contribute to the evolutionary pressure to maintain a catalytic Argonaute protein. The findings suggest that Ago2 catalysis is essential for miR-451 maturation and that this pathway is important for vertebrate development.