1. Plant Biology

Building customizable auto-luminescent luciferase-based reporters in plants

  1. Arjun Khakhar
  2. Colby G Starker
  3. James C Chamness
  4. Nayoung Lee
  5. Sydney Stokke
  6. Cecily Wang
  7. Ryan Swanson
  8. Furva Rizvi
  9. Takato Imaizumi
  10. Daniel F Voytas  Is a corresponding author
  1. University of Minnesota, United States
  2. University of Washington, United States
https://doi.org/10.7554/eLife.52786
Share this article
Cite this article
  1. Arjun Khakhar
  2. Colby G Starker
  3. James C Chamness
  4. Nayoung Lee
  5. Sydney Stokke
  6. Cecily Wang
  7. Ryan Swanson
  8. Furva Rizvi
  9. Takato Imaizumi
  10. Daniel F Voytas
(2020)
Building customizable auto-luminescent luciferase-based reporters in plants
eLife 9:e52786.
https://doi.org/10.7554/eLife.52786
  2. Download BibTeX
  3. Download .RIS

Abstract

Bioluminescence is a powerful biological signal that scientists have repurposed as a reporter for gene expression in plants and animals. However, there are downsides associated with the need to provide a substrate to these reporters, including its high cost and non-uniform tissue penetration. In this work we reconstitute a fungal bioluminescence pathway (FBP) in planta using a composable toolbox of parts. We demonstrate that the FBP can create luminescence across various tissues in a broad range of plants without external substrate addition. We also show how our toolbox can be used to deploy the FBP in planta to build auto-luminescent reporters for the study of gene-expression and hormone fluxes. A low-cost imaging platform for gene expression profiling is also described. These experiments lay the groundwork for future construction of programmable auto-luminescent plant traits, such as light driven plant-pollinator interactions or light emitting plant-based sensors.

Data availability

All the data collected for this study is depicted in the figures included in the manuscript. All the raw data used to make the figures has been uploaded.

Article and author information

Author details

  1. Arjun Khakhar

    Genetics, Cell Biology, and Development, University of Minnesota, Saint Paul, United States
    Competing interests
    The authors declare that no competing interests exist.

  2. Colby G Starker

    Genetics, Cell Biology, and Development, University of Minnesota, Saint Paul, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-6774-7227

  3. James C Chamness

    Genetics, Cell Biology, and Development, University of Minnesota, Saint Paul, United States
    Competing interests
    The authors declare that no competing interests exist.

  4. Nayoung Lee

    Biology, University of Washington, Seattle, United States
    Competing interests
    The authors declare that no competing interests exist.

  5. Sydney Stokke

    Genetics, Cell Biology, and Development, University of Minnesota, Saint Paul, United States
    Competing interests
    The authors declare that no competing interests exist.

  6. Cecily Wang

    Genetics, Cell Biology, and Development, University of Minnesota, Saint Paul, United States
    Competing interests
    The authors declare that no competing interests exist.

  7. Ryan Swanson

    Genetics, Cell Biology, and Development, University of Minnesota, Saint Paul, United States
    Competing interests
    The authors declare that no competing interests exist.

  8. Furva Rizvi

    Genetics, Cell Biology, and Development, University of Minnesota, Saint Paul, United States
    Competing interests
    The authors declare that no competing interests exist.

  9. Takato Imaizumi

    Biology, University of Washington, Seattle, United States
    Competing interests
    The authors declare that no competing interests exist.

  10. Daniel F Voytas

    Genetics, Cell Biology, and Development, University of Minnesota, Saint Paul, United States
    For correspondence
    voytas@umn.edu
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-4944-1224

Funding

University of Minnesota (Grand Challenges Postdoctoral Fellowship)

  • Arjun Khakhar

National Institutes of Health (R01GM079712)

  • Nayoung Lee

Rural Development Administration (PJ013386)

  • Nayoung Lee

U.S. Department of Energy

  • Colby G Starker
  • James C Chamness

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

Copyright

© 2020, Khakhar 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.

Metrics

  • 15,556
    views
  • 1,800
    downloads
  • 45
    citations

Views, downloads and citations are aggregated across all versions of this paper published by eLife.

Download links

A two-part list of links to download the article, or parts of the article, in various formats.

Downloads (link to download the article as PDF)

Open citations (links to open the citations from this article in various online reference manager services)

Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)

  1. Arjun Khakhar
  2. Colby G Starker
  3. James C Chamness
  4. Nayoung Lee
  5. Sydney Stokke
  6. Cecily Wang
  7. Ryan Swanson
  8. Furva Rizvi
  9. Takato Imaizumi
  10. Daniel F Voytas
(2020)
Building customizable auto-luminescent luciferase-based reporters in plants
eLife 9:e52786.
https://doi.org/10.7554/eLife.52786

Share this article

https://doi.org/10.7554/eLife.52786

Categories and tags

Research organism

Further reading

    1. Plant Biology

    Pectin methylesterase activity is required for RALF1 peptide signalling output

    Ann-Kathrin Rößling, Kai Dünser ... Jürgen Kleine-Vehn
    Research Article

    The extracellular matrix plays an integrative role in cellular responses in plants, but its contribution to the signalling of extracellular ligands largely remains to be explored. Rapid alkalinisation factors (RALFs) are extracellular peptide hormones that play pivotal roles in various physiological processes. Here, we address a crucial connection between the de-methylesterification machinery of the cell wall component pectin and RALF1 activity. Pectin is a polysaccharide, contributing to the structural integrity of the cell wall. Our data illustrate that the pharmacological and genetic interference with pectin methyl esterases (PMEs) abolishes RALF1-induced root growth repression. Our data suggest that positively charged RALF1 peptides bind negatively charged, de-methylesterified pectin with high avidity. We illustrate that the RALF1 association with de-methylesterified pectin is required for its FERONIA-dependent perception, contributing to the control of the extracellular matrix and the regulation of plasma membrane dynamics. Notably, this mode of action is independent of the FER-dependent extracellular matrix sensing mechanism provided by FER interaction with the leucine-rich repeat extensin (LRX) proteins. We propose that the methylation status of pectin acts as a contextualizing signalling scaffold for RALF peptides, linking extracellular matrix dynamics to peptide hormone-mediated responses.

    1. Biochemistry and Chemical Biology
    2. Plant Biology

    Structure and evolution of alanine/serine decarboxylases and the engineering of theanine production

    Hao Wang, Biying Zhu ... Zhaoliang Zhang
    Research Article

    Ethylamine (EA), the precursor of theanine biosynthesis, is synthesized from alanine decarboxylation by alanine decarboxylase (AlaDC) in tea plants. AlaDC evolves from serine decarboxylase (SerDC) through neofunctionalization and has lower catalytic activity. However, lacking structure information hinders the understanding of the evolution of substrate specificity and catalytic activity. In this study, we solved the X-ray crystal structures of AlaDC from Camellia sinensis (CsAlaDC) and SerDC from Arabidopsis thaliana (AtSerDC). Tyr341 of AtSerDC or the corresponding Tyr336 of CsAlaDC is essential for their enzymatic activity. Tyr111 of AtSerDC and the corresponding Phe106 of CsAlaDC determine their substrate specificity. Both CsAlaDC and AtSerDC have a distinctive zinc finger and have not been identified in any other Group II PLP-dependent amino acid decarboxylases. Based on the structural comparisons, we conducted a mutation screen of CsAlaDC. The results indicated that the mutation of L110F or P114A in the CsAlaDC dimerization interface significantly improved the catalytic activity by 110% and 59%, respectively. Combining a double mutant of CsAlaDCL110F/P114A with theanine synthetase increased theanine production 672% in an in vitro system. This study provides the structural basis for the substrate selectivity and catalytic activity of CsAlaDC and AtSerDC and provides a route to more efficient biosynthesis of theanine.