Trafficking of siRNA precursors by the dsRBD protein Blanks in Drosophila

Nucleic Acids Res. 2020 Apr 17;48(7):3906-3921. doi: 10.1093/nar/gkaa072.

Abstract

RNA interference targets aberrant transcripts with cognate small interfering RNAs, which derive from double-stranded RNA precursors. Several functional screens have identified Drosophila blanks/lump (CG10630) as a facilitator of RNAi, yet its molecular function has remained unknown. The protein carries two dsRNA binding domains (dsRBD) and blanks mutant males have a spermatogenesis defect. We demonstrate that blanks selectively boosts RNAi triggered by dsRNA of nuclear origin. Blanks binds dsRNA via its second dsRBD in vitro, shuttles between nucleus and cytoplasm and the abundance of siRNAs arising at many sites of convergent transcription is reduced in blanks mutants. Since features of nascent RNAs - such as introns and transcription beyond the polyA site - contribute to the small RNA pool, we propose that Blanks binds dsRNA formed by cognate nascent RNAs in the nucleus and fosters its export to the cytoplasm for dicing. We refer to the resulting small RNAs as blanks exported siRNAs (bepsiRNAs). While bepsiRNAs were fully dependent on RNA binding to the second dsRBD of blanks in transgenic flies, male fertility was not. This is consistent with a previous report that linked fertility to the first dsRBD of Blanks. The role of blanks in spermatogenesis appears thus unrelated to its role in dsRNA export.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Animals
  • Argonaute Proteins / metabolism
  • Cell Line
  • Cell Nucleus / metabolism
  • Drosophila / genetics
  • Drosophila / metabolism
  • Drosophila Proteins / chemistry
  • Drosophila Proteins / metabolism*
  • Fertility / genetics
  • Interspersed Repetitive Sequences
  • Male
  • Mutation
  • Protein Domains
  • RNA Helicases / metabolism
  • RNA Interference
  • RNA Precursors / metabolism*
  • RNA Transport
  • RNA, Antisense
  • RNA, Double-Stranded / metabolism
  • RNA, Small Interfering / metabolism*
  • RNA-Binding Proteins / chemistry
  • RNA-Binding Proteins / metabolism*
  • Ribonuclease III / metabolism
  • Transcription, Genetic

Substances

  • AGO2 protein, Drosophila
  • Argonaute Proteins
  • Drosophila Proteins
  • RNA Precursors
  • RNA, Antisense
  • RNA, Double-Stranded
  • RNA, Small Interfering
  • RNA-Binding Proteins
  • blanks protein, Drosophila
  • DCR-2 protein, Drosophila
  • Ribonuclease III
  • RNA Helicases