CRISPR/Cas9 and Active Genetics-based trans-species replacement of the endogenous Drosophila kni-L2 CRM reveals unexpected complexity

  1. Xiang-Ru Shannon Xu
  2. Valentino Matteo Gantz
  3. Natalia Siomava
  4. Ethan Bier  Is a corresponding author
  1. University of California, San Diego, United States
  2. University of Göttingen, Germany

Abstract

The knirps (kni) locus encodes transcription factors required for induction of the L2 wing vein in Drosophila. Here, we employ diverse CRISPR/Cas9 genome editing tools to generate a series of targeted lesions within the endogenous cis-regulatory module (CRM) required for kni expression in the L2 vein primordium. Phenotypic analysis of these 'in locus' mutations based on both expression of Kni protein and adult wing phenotypes, reveals novel unexpected features of L2-CRM function including evidence for a chromosome pairing-dependent process that promotes transcription. We also demonstrate that self-propagating active genetic elements (CopyCat elements) can efficiently delete and replace the L2-CRM with orthologous sequences from other divergent fly species. Wing vein phenotypes resulting from these trans-species enhancer replacements parallel features of the respective donor fly species. This highly sensitive phenotypic readout of enhancer function in a native genomic context reveals novel features of CRM function undetected by traditional reporter gene analysis.

Article and author information

Author details

  1. Xiang-Ru Shannon Xu

    Section of Cell and Developmental Biology, University of California, San Diego, La Jolla, United States
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-2781-9767
  2. Valentino Matteo Gantz

    Section of Cell and Developmental Biology, University of California, San Diego, La Jolla, United States
    Competing interests
    Valentino Matteo Gantz, Founder of Synbal, Inc. and Agragene, Inc.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-2453-0711
  3. Natalia Siomava

    Department of Developmental Biology, University of Göttingen, Göttingen, Germany
    Competing interests
    No competing interests declared.
  4. Ethan Bier

    Section of Cell and Developmental Biology, University of California, San Diego, La Jolla, United States
    For correspondence
    ebier@ucsd.edu
    Competing interests
    Ethan Bier, Founder of Synbal, Inc. and Agragene, Inc.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-2892-3005

Funding

National Institutes of Health (1R01GM117321)

  • Ethan Bier

Allen Foundation (Distinguished Investigator Award)

  • Ethan Bier

National Institutes of Health (1DP5OD023098)

  • Valentino Matteo Gantz

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

Reviewing Editor

  1. Nipam H Patel, University of California, Berkeley, United States

Version history

  1. Received: July 8, 2017
  2. Accepted: December 21, 2017
  3. Accepted Manuscript published: December 23, 2017 (version 1)
  4. Version of Record published: February 6, 2018 (version 2)
  5. Version of Record updated: February 28, 2018 (version 3)
  6. Version of Record updated: March 2, 2018 (version 4)

Copyright

© 2017, Xu 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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  1. Xiang-Ru Shannon Xu
  2. Valentino Matteo Gantz
  3. Natalia Siomava
  4. Ethan Bier
(2017)
CRISPR/Cas9 and Active Genetics-based trans-species replacement of the endogenous Drosophila kni-L2 CRM reveals unexpected complexity
eLife 6:e30281.
https://doi.org/10.7554/eLife.30281

Share this article

https://doi.org/10.7554/eLife.30281

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