Welcome to the new version of CaltechAUTHORS. Login is currently restricted to library staff. If you notice any issues, please email coda@library.caltech.edu
Published July 2011 | Supplemental Material + Published
Journal Article Open

Engineering ligand-responsive RNA controllers in yeast through the assembly of RNase III tuning modules


The programming of cellular networks to achieve new biological functions depends on the development of genetic tools that link the presence of a molecular signal to gene-regulatory activity. Recently, a set of engineered RNA controllers was described that enabled predictable tuning of gene expression in the yeast Saccharomyces cerevisiae through directed cleavage of transcripts by an RNase III enzyme, Rnt1p. Here, we describe a strategy for building a new class of RNA sensing-actuation devices based on direct integration of RNA aptamers into a region of the Rnt1p hairpin that modulates Rnt1p cleavage rates. We demonstrate that ligand binding to the integrated aptamer domain is associated with a structural change sufficient to inhibit Rnt1p processing. Three tuning strategies based on the incorporation of different functional modules into the Rnt1p switch platform were demonstrated to optimize switch dynamics and ligand responsiveness. We further demonstrated that these tuning modules can be implemented combinatorially in a predictable manner to further improve the regulatory response properties of the switch. The modularity and tunability of the Rnt1p switch platform will allow for rapid optimization and tailoring of this gene control device, thus providing a useful tool for the design of complex genetic networks in yeast.

Additional Information

© 2011 The Author(s). Published by Oxford University Press. This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/ by-nc/2.5), which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited. Received December 29, 2010; Revised January 27, 2011; Accepted February 3, 2011. First published online: February 25, 2011. We thank K. Hoff for assistance in the expression and purification of Rnt1p; and S. Bastian and F.H. Arnold for assistance in sonication and FPLC. The National Science Foundation (CAREER award CBET-0917705 to C.D.S.); Alfred P. Sloan Foundation, fellowship (to C.D.S.). Funding for open access charge: National Science Foundation CBET-0917705.

Attached Files

Published - Babiskin2011p15352Nucleic_Acids_Res.pdf

Supplemental Material - Supplementary_Information.pdf


Files (5.4 MB)
Name Size Download all
314.0 kB Preview Download
5.1 MB Preview Download

Additional details

August 19, 2023
October 23, 2023