Building customizable auto-luminescent luciferase-based reporters in plants
- University of Minnesota, United States
- University of Washington, United States
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
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.
Reviewing Editor
- Richard Amasino, University of Wisconsin Madison, United States
Publication history
- Received: October 16, 2019
- Accepted: March 24, 2020
- Accepted Manuscript published: March 25, 2020 (version 1)
- Version of Record published: April 17, 2020 (version 2)
- Version of Record updated: July 14, 2020 (version 3)
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.
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Building customizable auto-luminescent luciferase-based reporters in plants
eLife 9:e52786.
https://doi.org/10.7554/eLife.52786
Further reading
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- Plant Biology
A subset of plant intracellular NLR immune receptors detect effector proteins, secreted by phytopathogens to promote infection, through unconventional integrated domains which resemble the effector’s host targets. Direct binding of effectors to these integrated domains activates plant defenses. The rice NLR receptor Pik-1 binds the Magnaporthe oryzae effector AVR-Pik through an integrated heavy metal-associated (HMA) domain. However, the stealthy alleles AVR-PikC and AVR-PikF avoid interaction with Pik-HMA and evade host defenses. Here, we exploited knowledge of the biochemical interactions between AVR-Pik and its host target, OsHIPP19, to engineer novel Pik-1 variants that respond to AVR-PikC/F. First, we exchanged the HMA domain of Pikp-1 for OsHIPP19-HMA, demonstrating that effector targets can be incorporated into NLR receptors to provide novel recognition profiles. Second, we used the structure of OsHIPP19-HMA to guide the mutagenesis of Pikp-HMA to expand its recognition profile. We demonstrate that the extended recognition profiles of engineered Pikp-1 variants correlate with effector binding in planta and in vitro, and with the gain of new contacts across the effector/HMA interface. Crucially, transgenic rice producing the engineered Pikp-1 variants was resistant to blast fungus isolates carrying AVR-PikC or AVR-PikF. These results demonstrate that effector target-guided engineering of NLR receptors can provide new-to-nature disease resistance in crops.
