Artificial Repeat-Structured siRNA Precursors as Tunable Regulators for Saccharomyces cerevisiae.

RNA interference (RNAi) is widely used as a research tool for studying biological systems and implementing artificial genetic circuits that function by modulating RNA concentrations. Here we engineered Saccharomyces cerevisiae containing a heterologous Saccharomyces castelli RNAi system as a test-bed for RNAi-based circuits. Unlike prior approaches, we describe a strategy that leverages repeat-structured siRNA precursors with incrementally sized stems formed from 23 bp-repeats to achieve modular RNAi-based gene regulation. These enable repression strength to be tuned in a systematic manner by changing the size of the siRNA precursor hairpin stem, without modifying the number or sequence of target sites in the target RNA. We demonstrate that this hairpin-based regulation is able to target both cytoplasmic and nuclear localized RNAs and is stable over extended growth periods. This platform enables the targeting of cellular RNAs as a tunable regulatory layer for sophisticated gene circuits in Saccharomyces cerevisiae.