Development of a synthetic molecular sensor for oxygen in plants

The use of genetically encoded biosensors to selectively track molecules of interest holds great potential in understanding how nutrients, hormones and other signaling molecules relocate in response to environmental and developmental cues. Among these, oxygen availability represents an essential cue to drive biological processes, due to the involvement of oxygen as a substrate in many metabolic reactions. Indeed, the need to attune metabolic and developmental processes to oxygen availability promoted the evolution of different sensing strategies. In higher plants, this was achieved with oxygen-dependent proteolysis of specific ethylene responsive factors (ERF-VII). In the frame of my PhD, I first investigated whether protein motifs conserved among these transcription factors contribute to DNA binding. Moreover, I selected signaling modules across eukaryotic kingdoms to design and optimize an orthogonal oxygen-responsive device to be expressed in plants. This was achieved through rational combinations of mammalian, plant and yeast protein domains, to generate a series of genetic circuits that generate alternative outputs in an oxygen-dependent manner. After extensive characterization of these synthetic devices by means of transient expression in plant cells, I stably introduced them in the Arabidopsis thaliana genome to evaluate internal oxygen dynamics and promote advantageous developmental strategies in response to hypoxic conditions. In summary, the work presented here represents a pioneering effort to combine protein features from different organisms to engineer novel biological functions. It also provides an initial exploration of possible applications for molecular switches based on oxygen availability and paves the way towards its deployment in different scientific fields.