Abstract
Optogenetics is the genetic approach for controlling cellular processes with light. It provides spatiotemporal, quantitative and reversible control over biological signaling and metabolic processes, overcoming limitations of chemically inducible systems. However, optogenetics lags in plant research because ambient light required for growth leads to undesired system activation. We solved this issue by developing plant usable light-switch elements (PULSE), an optogenetic tool for reversibly controlling gene expression in plants under ambient light. PULSE combines a blue-light-regulated repressor with a red-light-inducible switch. Gene expression is only activated under red light and remains inactive under white light or in darkness. Supported by a quantitative mathematical model, we characterized PULSE in protoplasts and achieved high induction rates, and we combined it with CRISPR-Cas9-based technologies to target synthetic signaling and developmental pathways. We applied PULSE to control immune responses in plant leaves and generated Arabidopsis transgenic plants. PULSE opens broad experimental avenues in plant research and biotechnology.
Highlights
The reversible and orthogonal control of cellular processes with high spatiotemporal resolution is key for quantitatively understanding the dynamics of biological signalling networks as well as for programming desired phenotypes
Due to the spectral compatibility limitations described above or the need for co-factors difficult to administer to whole plants, these tools could only be applied in transiently transformed plant cells such as mesophyll protoplasts from Nicotiana tabacum or Arabidopsis thaliana, and the moss
PULSE is an integrated optogenetic molecular device, consisting of two components, a module providing activation of gene expression under red light (ROn) and a second one ensuring effective transcriptional repression under blue light (BOff) (Fig. 1). The rationale behind this new conceptual and experimental approach is that the combination of both switches will yield a system that is inactive in ambient growth conditions and only active upon irradiation with red light
Summary
The reversible and orthogonal control of cellular processes with high spatiotemporal resolution is key for quantitatively understanding the dynamics of biological signalling networks as well as for programming desired phenotypes. We set here to develop the first optogenetic system for the control of gene expression in plants that is silent under white light and can be activated with monochromatic red light.
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