Reprogramming the piRNA pathway for multiplexed and transgenerational gene silencing in C. elegans

Monika Priyadarshini, Julie Zhouli Ni, Amhed M. Vargas-Velazquez, Sam Guoping Gu, Christian Frøkjær-Jensen

Research output: Contribution to journalArticlepeer-review

12 Scopus citations

Abstract

Single-guide RNAs can target exogenous CRISPR-Cas proteins to unique DNA locations, enabling genetic tools that are efficient, specific and scalable. Here we show that short synthetic guide Piwi-interacting RNAs (piRNAs) (21-nucleotide sg-piRNAs) expressed from extrachromosomal transgenes can, analogously, reprogram the endogenous piRNA pathway for gene-specific silencing in the hermaphrodite germline, sperm and embryos of Caenorhabditis elegans. piRNA-mediated interference ('piRNAi') is more efficient than RNAi and can be multiplexed, and auxin-mediated degradation of the piRNA-specific Argonaute PRG-1 allows conditional gene silencing. Target-specific silencing results in decreased messenger RNA levels, amplification of secondary small interfering RNAs and repressive chromatin modifications. Short (300 base pairs) piRNAi transgenes amplified from arrayed oligonucleotide pools also induce silencing, potentially making piRNAi highly scalable. We show that piRNAi can induce transgenerational epigenetic silencing of two endogenous genes (him-5 and him-8). Silencing is inherited for four to six generations after target-specific sg-piRNAs are lost, whereas depleting PRG-1 leads to essentially permanent epigenetic silencing.
Original languageEnglish (US)
JournalNature Methods
DOIs
StatePublished - Feb 3 2022

Bibliographical note

KAUST Repository Item: Exported on 2022-02-07
Acknowledged KAUST grant number(s): CRG, OSR
Acknowledgements: We thank L. Wahba and A. Fire (Stanford University) for sharing reagents before publication. Some strains were provided by the CGC, which is funded by the NIH Office of Research Infrastructure Programs (grant no. P40 OD010440). The research was funded by KAUST Office of Sponsored Research grant no. OSR-CRG2019-4016 (C.F.-J.), a Rutgers University Busch Biomedical Grant (S.G.G.) and the National Institute of General Medical Sciences of the National Institutes of Health under award number R01GM111752 (S.G.G.). The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.

ASJC Scopus subject areas

  • Biochemistry
  • Cell Biology
  • Molecular Biology
  • Biotechnology

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