title: Functional small RNAs are generated from select miRNA hairpin loops in flies and mammals.
abstract: In the canonical animal microRNA (miRNA) pathway, Drosha generates ∼60- to 70-nucleotide pre-miRNA hairpins that are cleaved by Dicer into small RNA duplexes that load into Argonaute proteins, which retain a single mature strand in the active complex. The terminal loops of some miRNA hairpins regulate processing efficiency, but once liberated by Dicer, they are generally considered nonfunctional by-products. Here, we show that specific miRNA loops accumulate in effector Argonaute complexes in Drosophila and mediate miRNA-type repression. This was unexpected, since endogenous loading of Argonaute proteins was believed to occur exclusively via small RNA duplexes. Using in vitro assays, which recapitulate Argonaute-specific loop loading from synthetic pre-miRNAs and even single-stranded oligoribonucleotides corresponding to miRNA loops, we reveal that the loop-loading mechanism is distinct from duplex loading. We also show that miRNA loops loaded into the miRNA effector AGO1 are subject to 3' resection, and structure-function analyses indicate selectivity of loop loading. Finally, we demonstrate that select miRNA loops in mammals are similarly loaded into Argonaute complexes and direct target repression. Altogether, we reveal a conserved mechanism that yields functional RNAs from miRNA loop regions, broadening the repertoire of Argonaute-dependent regulatory RNAs and providing evidence for functionality of endogenous ssRNA species.
authors.affiliation: Department of Developmental Biology, Sloan-Kettering Institute, New York, New York 10065, USA. okamurak@tll.org.sg
authors.forename: Katsutomo
authors.initials: K
authors.lastname: Okamura
authors.forename: Erik
authors.initials: E
authors.lastname: Ladewig
authors.forename: Li
authors.initials: L
authors.lastname: Zhou
authors.forename: Eric C
authors.initials: EC
authors.lastname: Lai
mesh_terms: D000818:Animals; D060565:Argonaute Proteins / Q000378:metabolism; D017124:Conserved Sequence; D029721:Drosophila Proteins / Q000378:metabolism; D004331:Drosophila melanogaster / Q000235:genetics* / Q000378:metabolism*; D005786:Gene Expression Regulation; D055106:Genome-Wide Association Study; D008322:Mammals; D051379:Mice; D035683:MicroRNAs / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D009690:Nucleic Acid Conformation
chemical_list: C489654:AGO1 protein, Drosophila; C433807:AGO2 protein, Drosophila; D060565:Argonaute Proteins; D029721:Drosophila Proteins; D035683:MicroRNAs
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