An inducible system inspired in the bacterial quórum sensing mechanism for antimicrobial protein expression in Solanaceae plastids

SEGRETIN ME; BOCCARDO NA; BRAVO ALMONACID FF

Rhodas

Congreso; XVI International Congress on Molecular Plant-Microbe Interactions; 2014

Plant diseases affect most staple crops decreasing productivity worldwide and severely compromising food security. Many attempts have been done to obtain disease resistant crops through transgenesis, with variable results. One strategy is to express heterologous proteins with antimicrobial activity. For this strategy to be successful, high levels of protein accumulation are required, although this could result in lession-mimic phenotypes. In order to overcome these limitations, we decided to evaluate plastid transformation as an alternative strategy. Plastome transformation advantages nuclear transformation because of the higher genome copy number, absence of positional effects and silencing, and maternal inheritance of transgenes. We developed a plastid transformation vector and used it to transform N. tabacum plastids with three antimicrobial proteins, alone or combined as polycistrons: tobacco PR proteins AP24 and -1,3-glucanase, and dermaseptin S1 from Phyllomedusa sauvagii. We obtained transplastomic plants that are being characterized. In order to avoid drawbacks due to constitutive high expression levels of antimicrobial proteins, we are developing an inducible expression system inspired in the bacterial Quorum Sensing (QS) mechanism. We assembled in a plastid transformation vector the following QS elements from Pseudomonas aeruginosa: lasB promoter sequence was cloned upstream uidA reporter gene, and CDS corresponding to regulatory proteins lasR and RhlR were included alone or combined as a polycistron downstream psbA promoter sequence. Transplastomic plants were obtained and are being assessed for GUS expression after treatment with C4-HSL and 3-oxo-C12-HSL homoserin lactones (HSL), the signalling molecules that together with their cognate regulatory proteins (lasR and RhlR, respectively) allow transcription from lasB promoter.