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MAEL-dependent selective processing of pachytene piRNA precursors and translation of spermiogenic mRNAs

ABSTRACT: Piwi-interacting small RNAs (piRNAs) of fetal prospermatogonia of mice have been strongly implicated in transposon control. In contrast, little is known about biogenesis and function of abundant piRNAs from adult testes expressed in late spermatocytes and round spermatids. These so-called "pachytene" piRNAs are processed from long non-coding piRNA precursors and have no defined RNA targets in the transcriptome even though their binding partner Piwi, MIWI, is essential for spermiogenesis and fertility. Here we report that 129SvJae mice lacking Maelstrom (MAEL), a conserved piRNA pathway protein, exhibit spermiogenic arrest with defects in acrosome and flagellum formation. Further analysis revealed MAEL association with RNPs containing MIWI, TDRD6, and processed intermediates of pachytene piRNA precursors of various length. Loss of MAEL causes a 10-fold drop in pachytene piRNA levels but an increase in piRNAs from abundantly expressed mRNAs. These results suggest a MAEL-dependent mechanism for the selective processing of pachytene piRNA precursor into piRNAs. Strikingly, ribosome profiling of Mael-null testes revealed that reduced piRNA production is accompanied by reduced translation of over 800 spermiogenic mRNAs including those encoding acrosome and flagellum proteins. In light of recent reports of piRNA-independent protection of translationally repressed mRNPs by MIWI and piRNA-dependent turnover of MIWI, we propose that pachytene piRNAs function by controlling the availably of MIWI for the translational repression of spermiogenic mRNAs.

ORGANISM(S): Mus musculus

PROVIDER: GSE50983 | GEO | 2014/07/25

SECONDARY ACCESSION(S): PRJNA219480

REPOSITORIES: GEO

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Project description:Pachytene piRNAs are PIWI-interacting small RNAs abundantly expressed in pachytene spermatocytes and spermatids in adult mouse testes. Both MIWI and MILI-bound pachytene piRNAs have been found enriched in round spermatids. Miwi-null male mice are sterile due to spermiogenic arrest. In C. elegans, sperm-borne piRNAs appear to have an epigenetic role during fertilization and development because progeny of offspring derived from piRNA-deficient sperm display a progressive fertility loss after several generations. In mice, it remains unknown whether MIWI-bound pachytene piRNA-deficient round spermatids can produce offspring, and whether the progeny of offspring derived from MIWI-bound pachytene piRNA-deficient round spermatids also exhibit transgenerational loss of fertility. Here, we report that Miwi KO round spermatids could fertilize both wild type (WT) and Miwi KO oocytes through round spermatid microinjection (ROSI), and produce healthy and fertile offspring despite the aberrant pachytene piRNA profiles in those Miwi KO spermatids. Progeny of ROSI-derived heterozygotes, both male and female, displayed normal fertility for at least three generations when bred with either WT or Miwi KO females. Our data indicate that aberrant MIWI-bound pachytene piRNAs profiles in spermatids do not affect fertilization, early embryonic development, or fertility of the offspring, suggesting a normal pachytene piRNAs profile is not required for paternal transgenerational epigenetic inheritance in mice. Method: Round spermatids were purified from WT and Miwi KO adult testes using a mini-STA-PUT method[Methods Enzymol 1993; 225:84-113.]，the purity of round spermatids was >90% based on our previous report[ J Biol Chem 2012; 287:25173-25190.]. Small RNA was isolated from round spermatids using the mirVana RNA isolation kit (Ambion) according to the manufacturer’s instructions. RNA quality and quantity were assessed using the Agilent 2100 Bioanalyzer. Small RNA-Seq was performed on an Ion Proton sequencer (Life Technologies). Libraries were prepared using the Ion Total RNA-Seq Kit v2 (Invitrogen) with biological triplicates for WT and Miwi KO samples. Resutls:Our data indicate that aberrant MIWI-bound pachytene piRNAs profiles in spermatids do not affect fertilization, early embryonic development, or fertility of the offspring, suggesting a normal pachytene piRNAs profile is not required for paternal transgenerational epigenetic inheritance in mice.

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MAEL-dependent selective processing of pachytene piRNA precursors and translation of spermiogenic mRNAs

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