A novel biosensor to monitor proline in pea root exudates and nodules under osmotic stress and recovery

Abstract

Background and aims Plant and bacteria are able to synthesise proline, which acts as a compound to counteract the negative effects of osmotic stresses. Most methodologies rely on the extraction of compounds using destructive methods. This work describes a new proline biosensor that allows the monitoring of proline levels in a non-invasive manner in root exudates and nodules of legume plants. Methods The proline biosensor was constructed by cloning the promoter region of pRL120553, a gene with high levels of induction in the presence of proline, in front of the lux cassette in Rhizobium leguminosarum bv. viciae. Results Free-living assays show that the proline biosensor is sensitive and specific for proline. Proline was detected in both root exudates and nodules of pea plants. The luminescence detected in bacteroids did not show variations during osmotic stress treatments, but significantly increased during recovery. Conclusions This biosensor is a useful tool for the in vivo monitoring of proline levels in root exudates and bacteroids of symbiotic root nodules, and it contributes to our understanding of the metabolic exchange occurring in nodules under abiotic stress conditions.

Transcriptomic Responses of Rhizobium phaseoli to Root Exudates Reflect Its Capacity to Colonize Maize and Common Bean in an Intercropping System

“…Proline is an abundant amino acid in maize exudates (Phillips et al, 2004) and is also produced by bean roots at different growth steps (Tawaraya et al, 2014). The putA gene has been used as a biosensor to monitor the exudation of proline by pea roots (Rubia et al, 2020) and its overexpression in Sinorhizobium meliloti strains increases their competitiveness to colonize and nodule alfalfa (Medicago sativa L.; van Dillewijn et al, 2001). In this work, putA expression was induced in an exudate exposure of 2 h, but no statistically significant differences were observed between plant treatments (Figure 6).…”

Modern analytical and bioanalytical technologies and concepts for smart and precision farming

“…Genetic engineering of biorecognition molecules could be a strategy to improve the performance of biosensors. The fusion of reporter genes such as green fluorescent protein (GFP), firefly luciferase (luc), and bacterial luciferase (lux) with microorganisms such as bacteria 14 and fungi 15 increases the sensitivity and allows imaging purposes. The biological response can be converted to measurable luminescence and fluorescence signals, enabling quantitation analysis.…”

“…52 An optical in vivo biosensor made of genetically engineered bacteria, Rhizobium leguminosarum , was developed to detect osmotic pressure by measuring proline levels in the root. 14 The bacterium was genetically engineered with a plasmid consisting of the promoter pRL120553 and the lux operon. Luminescence changes that vary according to the proline level will reflect the osmotic pressure experienced by the plant.…”

Optical Sensing Technologies to Elucidate the Interplay between Plant and Microbes

“…Proline concentrations in pea root exudates under water-stress conditions were detected using a lux biosensor. The biosensor was constructed by cloning the promoter sequence of pRL120553 [ 189 ].…”