ABSTRACT
Enhancer RNAs (eRNAs) are long non-coding RNAs that originate from enhancers. Although eRNA transcription is a canonical feature of activated enhancers, the molecular features required for eRNA function and the mechanism of how eRNAs impinge on target gene transcription have not been established. Thus, using eRNA-dependent RNA polymerase II (Pol II) pause release as a model, we here investigate the requirement of sequence, structure and length of eRNAs for their ability to stimulate Pol II pause release by detaching NELF from paused Pol II. We find eRNAs not to exert their function through common structural or sequence motifs. Instead, eRNAs that exhibit a length >200 nucleotides and that contain unpaired guanosines make multiple, allosteric contacts with NELF subunits -A and -E to trigger efficient NELF release. By revealing the molecular determinants of eRNA function, our study establishes eRNAs as an important player in Pol II pause release, and it provides new insight into the regulation of metazoan transcription.
Subject(s)
RNA Polymerase II , RNA, Long Noncoding , Animals , Enhancer Elements, Genetic , Gene Expression Regulation , Promoter Regions, Genetic , RNA Polymerase II/metabolism , RNA, Long Noncoding/physiology , Regulatory Sequences, Nucleic Acid , Transcription, GeneticABSTRACT
PIWI proteins utilize small RNAs called piRNAs to silence transposable elements, thereby protecting germline integrity. In planarian flatworms, PIWI proteins are essential for regeneration, which requires adult stem cells termed neoblasts. Here, we characterize planarian piRNAs and examine the roles of PIWI proteins in neoblast biology. We find that the planarian PIWI proteins SMEDWI-2 and SMEDWI-3 cooperate to degrade active transposons via the ping-pong cycle. Unexpectedly, we discover that SMEDWI-3 plays an additional role in planarian mRNA surveillance. While SMEDWI-3 degrades numerous neoblast mRNAs in a homotypic ping-pong cycle, it is also guided to another subset of neoblast mRNAs by antisense piRNAs and binds these without degrading them. Mechanistically, the distinct activities of SMEDWI-3 are primarily dictated by the degree of complementarity between target mRNAs and antisense piRNAs. Thus, PIWI proteins enable planarians to repurpose piRNAs for potentially critical roles in neoblast mRNA turnover.
