1. Biochemistry and Chemical Biology
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Rational design of aptazyme riboswitches for efficient control of gene expression in mammalian cells

  1. Guocai Zhong  Is a corresponding author
  2. Haimin Wang
  3. Charles C Bailey
  4. Guangping Gao
  5. Michael Farzan  Is a corresponding author
  1. The Scripps Research Institute, United States
  2. Johns Hopkins School of Medicine, United States
  3. University of Massachusetts Medical School, United States
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Cite this article as: eLife 2016;5:e18858 doi: 10.7554/eLife.18858

Abstract

Efforts to control mammalian gene expression with ligand-responsive riboswitches have been hindered by lack of a general method for generating efficient switches in mammalian systems. Here we describe a rational-design approach that enables rapid development of efficient cis-acting aptazyme riboswitches. We identified communication-module characteristics associated with aptazyme functionality through analysis of a 32-aptazyme test panel. We then developed a scoring system that predicts an aptazymes's activity by integrating three characteristics of communication-module bases: hydrogen bonding, base stacking, and distance to the enzymatic core. We validated the power and generality of this approach by designing aptazymes responsive to three distinct ligands, each with markedly wider dynamic ranges than any previously reported. These aptayzmes efficiently regulated adeno-associated virus (AAV)-vectored transgene expression in cultured mammalian cells and mice, highlighting one application of these broadly usable regulatory switches. Our approach enables efficient, protein-independent control of gene expression by a range of small molecules.

Article and author information

Author details

  1. Guocai Zhong

    Department of Immunology and Microbial Sciences, The Scripps Research Institute, Jupiter, United States
    For correspondence
    gzhong@scripps.edu
    Competing interests
    The authors declare that no competing interests exist.
  2. Haimin Wang

    Department of Immunology and Microbial Sciences, The Scripps Research Institute, Jupiter, United States
    Competing interests
    The authors declare that no competing interests exist.
  3. Charles C Bailey

    Department of Molecular and Comparative Pathology, Johns Hopkins School of Medicine, Baltimore, United States
    Competing interests
    The authors declare that no competing interests exist.
  4. Guangping Gao

    Gene Therapy Center, University of Massachusetts Medical School, Worcester, United States
    Competing interests
    The authors declare that no competing interests exist.
  5. Michael Farzan

    Department of Immunology and Microbial Sciences, The Scripps Research Institute, Jupiter, United States
    For correspondence
    mfarzan@scripps.edu
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-2990-5319

Funding

National Institutes of Health (R01 AI091476)

  • Michael Farzan

National Institutes of Health (P01 AI100263)

  • Michael Farzan

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Ethics

Animal experimentation: This study was performed in strict accordance with the recommendations in the Guide for the Care and Use of Laboratory Animals of the National Institutes of Health. All of the animals were handled according to approved institutional animal care and use committee (IACUC) protocol (#14-028) of the Scripps Florida.

Reviewing Editor

  1. Ronald Breaker, Yale University, United States

Publication history

  1. Received: June 15, 2016
  2. Accepted: November 1, 2016
  3. Accepted Manuscript published: November 2, 2016 (version 1)
  4. Version of Record published: November 30, 2016 (version 2)

Copyright

© 2016, Zhong et al.

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.

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