Abstract
Short regulatory RNA molecules underpin gene expression and govern cellular state and physiology. To establish an alternative layer of control over these processes, we generated chimeric regulatory RNAs that interact reversibly and light-dependently with the light-oxygen-voltage photoreceptor PAL. By harnessing this interaction, the function of micro RNAs (miRs) and short hairpin (sh) RNAs in mammalian cells can be regulated in a spatiotemporally precise manner. The underlying strategy is generic and can be adapted to near-arbitrary target sequences. Owing to full genetic encodability, it establishes optoribogenetic control of cell state and physiology. The method stands to facilitate the non-invasive, reversible and spatiotemporally resolved study of regulatory RNAs and protein function in cellular and organismal environments.
Highlights
Short regulatory RNA molecules underpin gene expression and govern cellular state and physiology
We first generated aptamer-modified variants of premiR-21 (SHA, Fig. 2a) and analyzed them in reporter gene assays that employ the expression of secreted Metrida luciferase or of enhanced green fluorescent protein with micro RNAs (miRs)-21 target sites embedded in the 3′-UTRs of the respective mRNA (Supplementary Fig. 1a, b, 2)[31]
By offering full genetic encodability, reversibility, and noninvasiveness combined with a small genetic footprint, our approach transcends previous approaches for controlling regulatory RNA activity
Summary
Short regulatory RNA molecules underpin gene expression and govern cellular state and physiology. To establish an alternative layer of control over these processes, we generated chimeric regulatory RNAs that interact reversibly and light-dependently with the lightoxygen-voltage photoreceptor PAL By harnessing this interaction, the function of micro RNAs (miRs) and short hairpin (sh) RNAs in mammalian cells can be regulated in a spatiotemporally precise manner. The chimeric RNAs enable the spatiotemporal control of short regulatory RNA function in mammalian cells, as we showcase for the lightdependent control of gene expression and cell-cycle progression This hitherto unavailable modality establishes a versatile RNA control system for analyzing various protein and miR functionalities in a reversible, spatiotemporally resolved, and non-invasive manner, and with full genetic encoding. Owing to the modularity of the chimeric RNAs, the technology readily applies to neararbitrary shRNAs
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