Tag Archives: piRNA

Mili and Miwi target RNA repertoire reveals piRNA biogenesis and function of Miwi in spermiogenesis.

Publication
MILI_MIWI

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Nat Struct Mol Biol. 19(8):773-81

Abstract
Germ cells implement elaborate mechanisms to protect their genetic material and to regulate gene expression during differentiation. Piwi proteins bind Piwi-interacting RNAs (piRNAs), small germline RNAs whose biogenesis and functions are still largely elusive. We used high-throughput sequencing after cross-linking and immunoprecipitation (HITS-CLIP) coupled with RNA-sequencing (RNA-seq) to characterize the genome-wide target RNA repertoire of Mili (Piwil2) and Miwi (Piwil1), two Piwi proteins expressed in mouse postnatal testis. We report the in vivo pathway of primary piRNA biogenesis and implicate distinct nucleolytic activities that process Piwi-bound precursor transcripts. Our studies indicate that pachytene piRNAs are the end products of RNA processing. HITS-CLIP demonstrated that Miwi binds spermiogenic mRNAs directly, without using piRNAs as guides, and independent biochemical analyses of testis mRNA ribonucleoproteins (mRNPs) established that Miwi functions in the formation of mRNP complexes that stabilize mRNAs essential for spermiogenesis.

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Mitochondrial protein BmPAPI modulates the length of mature piRNAs.

Publication
Papi

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RNA 19(10):1405-18

Abstract
PIWI proteins and their associated PIWI-interacting RNAs (piRNAs) protect genome integrity by silencing transposons in animal germlines. The molecular mechanisms and components responsible for piRNA biogenesis remain elusive. PIWI proteins contain conserved symmetrical dimethylarginines (sDMAs) that are specifically targeted by TUDOR domain-containing proteins. Here we report that the sDMAs of PIWI proteins play crucial roles in PIWI localization and piRNA biogenesis in Bombyx mori-derived BmN4 cells, which harbor fully functional piRNA biogenesis machinery. Moreover, RNAi screenings for Bombyx genes encoding TUDOR domain-containing proteins identified BmPAPI, a Bombyx homolog of Drosophila PAPI, as a factor modulating the length of mature piRNAs. BmPAPI specifically recognized sDMAs and interacted with PIWI proteins at the surface of the mitochondrial outer membrane. BmPAPI depletion resulted in 3′-terminal extensions of mature piRNAs without affecting the piRNA quantity. These results reveal the BmPAPI-involved piRNA precursor processing mechanism on mitochondrial outer membrane scaffolds.

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The RNA helicase MOV10L1 binds piRNA precursors to initiate piRNA processing

Publication
MOV10L1

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Genes Dev. 2015 Mar 15;29(6):617-29.

Abstract
Piwi-piRNA (Piwi-interacting RNA) ribonucleoproteins (piRNPs) enforce retrotransposon silencing, a function critical for preserving the genome integrity of germ cells. The molecular functions of most of the factors that have been genetically implicated in primary piRNA biogenesis are still elusive. Here we show that MOV10L1 exhibits 5′-to-3′ directional RNA-unwinding activity in vitro and that a point mutation that abolishes this activity causes a failure in primary piRNA biogenesis in vivo. We demonstrate that MOV10L1 selectively binds piRNA precursor transcripts and is essential for the generation of intermediate piRNA processing fragments that are subsequently loaded to Piwi proteins. Multiple analyses suggest an intimate coupling of piRNA precursor processing with elements of local secondary structures such as G quadruplexes. Our results support a model in which MOV10L1 RNA helicase activity promotes unwinding and funneling of the single-stranded piRNA precursor transcripts to the endonuclease that catalyzes the first cleavage step of piRNA processing.

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