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Mol Cell. 2019 Jul 9. pii: S1097-2765(19)30474-5. doi: 10.1016/j.molcel.2019.06.018. [Epub ahead of print]

Time-Resolved Small RNA Sequencing Unravels the Molecular Principles of MicroRNA Homeostasis.

Author information

1
Institute of Molecular Biotechnology (IMBA), Vienna BioCenter (VBC), 1030 Vienna, Austria.
2
Institute of Molecular Biotechnology (IMBA), Vienna BioCenter (VBC), 1030 Vienna, Austria. Electronic address: stefan.ameres@imba.oeaw.ac.at.

Abstract

Argonaute-bound microRNAs silence mRNA expression in a dynamic and regulated manner to control organismal development, physiology, and disease. We employed metabolic small RNA sequencing for a comprehensive view on intracellular microRNA kinetics in Drosophila. Based on absolute rate of biogenesis and decay, microRNAs rank among the fastest produced and longest-lived cellular transcripts, disposing up to 105 copies per cell at steady-state. Mature microRNAs are produced within minutes, revealing tight intracellular coupling of biogenesis that is selectively disrupted by pre-miRNA-uridylation. Control over Argonaute protein homeostasis generates a kinetic bottleneck that cooperates with non-coding RNA surveillance to ensure faithful microRNA loading. Finally, regulated small RNA decay enables the selective rapid turnover of Ago1-bound microRNAs, but not of Ago2-bound small interfering RNAs (siRNAs), reflecting key differences in the robustness of small RNA silencing pathways. Time-resolved small RNA sequencing opens new experimental avenues to deconvolute the timescales, molecular features, and regulation of small RNA silencing pathways in living cells.

KEYWORDS:

Argonaute; RNA expression dynamics; RNA metabolism; metabolic RNA labeling; microRNAs; post-transcriptional gene regulation; small RNA homeostasis; small RNA silencing; time-resolved RNA sequencing

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