Mili and Miwi Target RNA Repertoire Reveals piRNA Biogenesis and Function of Miwi in Spermiogenesis

Nat Struct Mol Biol. 2012 Aug;19(8):773-81. doi: 10.1038/nsmb.2347. Epub 2012 Jul 29.

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.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Argonaute Proteins / deficiency
  • Argonaute Proteins / genetics*
  • Argonaute Proteins / physiology*
  • Base Sequence
  • Gene Expression Regulation
  • Heterozygote
  • High-Throughput Nucleotide Sequencing
  • Immunoprecipitation
  • Male
  • Mice
  • Mice, Knockout
  • MicroRNAs / genetics
  • MicroRNAs / metabolism
  • Models, Biological
  • RNA Stability
  • RNA, Small Interfering / biosynthesis*
  • RNA, Small Interfering / genetics*
  • Ribonucleoproteins / genetics
  • Ribonucleoproteins / metabolism
  • Spermatogenesis / genetics*
  • Spermatogenesis / physiology*
  • Testis / metabolism

Substances

  • Argonaute Proteins
  • MicroRNAs
  • Piwil1 protein, mouse
  • Piwil2 protein, mouse
  • RNA, Small Interfering
  • Ribonucleoproteins
  • messenger ribonucleoprotein

Associated data

  • GEO/GSE27609
  • GEO/GSE27622