title: Programmable ligand detection system in plants through a synthetic signal transduction pathway.
abstract: BACKGROUND
  There is an unmet need to monitor human and natural environments for substances that are intentionally or unintentionally introduced. A long-sought goal is to adapt plants to sense and respond to specific substances for use as environmental monitors. Computationally re-designed periplasmic binding proteins (PBPs) provide a means to design highly sensitive and specific ligand sensing capabilities in receptors. Input from these proteins can be linked to gene expression through histidine kinase (HK) mediated signaling. Components of HK signaling systems are evolutionarily conserved between bacteria and plants. We previously reported that in response to cytokinin-mediated HK activation in plants, the bacterial response regulator PhoB translocates to the nucleus and activates transcription. Also, we previously described a plant visual response system, the de-greening circuit, a threshold sensitive reporter system that produces a visual response which is remotely detectable and quantifiable.
  
  
  RESULTS
  We describe assembly and function of a complete synthetic signal transduction pathway in plants that links input from computationally re-designed PBPs to a visual response. To sense extracellular ligands, we targeted the computational re-designed PBPs to the apoplast. PBPs bind the ligand and develop affinity for the extracellular domain of a chemotactic protein, Trg. We experimentally developed Trg fusions proteins, which bind the ligand-PBP complex, and activate intracellular PhoR, the HK cognate of PhoB. We then adapted Trg-PhoR fusions for function in plants showing that in the presence of an external ligand PhoB translocates to the nucleus and activates transcription. We linked this input to the de-greening circuit creating a detector plant.
  
  
  CONCLUSIONS
  Our system is modular and PBPs can theoretically be designed to bind most small molecules. Hence our system, with improvements, may allow plants to serve as a simple and inexpensive means to monitor human surroundings for substances such as pollutants, explosives, or chemical agents.
authors.affiliation: Department of Biology, Colorado State University, Fort Collins, Colorado, United States of America.
authors.forename: Mauricio S
authors.initials: MS
authors.lastname: Antunes
authors.forename: Kevin J
authors.initials: KJ
authors.lastname: Morey
authors.forename: J Jeff
authors.initials: JJ
authors.lastname: Smith
authors.forename: Kirk D
authors.initials: KD
authors.lastname: Albrecht
authors.forename: Tessa A
authors.initials: TA
authors.lastname: Bowen
authors.forename: Jeffrey K
authors.initials: JK
authors.lastname: Zdunek
authors.forename: Jared F
authors.initials: JF
authors.lastname: Troupe
authors.forename: Matthew J
authors.initials: MJ
authors.lastname: Cuneo
authors.forename: Colleen T
authors.initials: CT
authors.lastname: Webb
authors.forename: Homme W
authors.initials: HW
authors.lastname: Hellinga
authors.forename: June I
authors.initials: JI
authors.lastname: Medford
mesh_terms: D004784:Environmental Monitoring / Q000379:methods*; D018506:Gene Expression Regulation, Plant; D005818:Genetic Engineering; D000071677:Histidine Kinase; D008024:Ligands; D033902:Periplasmic Binding Proteins / Q000235:genetics* / Q000378:metabolism; D010944:Plants / Q000378:metabolism*; D011485:Protein Binding; D011494:Protein Kinases / Q000378:metabolism; D015398:Signal Transduction
chemical_list: D008024:Ligands; D033902:Periplasmic Binding Proteins; D011494:Protein Kinases; D000071677:Histidine Kinase