VOCs perception in Solanum lycopersicum: multiple approaches to a complex problem. Percezione dei VOCs in Solanum lycopersicum: diversi approcci per lo studio di un problema complesso

This doctoral thesis presents the background, methodologies, and results of the research project aimed at deepening our understanding of the mechanisms underlying the ability of plants to perceive volatile organic compounds (VOCs). Initially, the effect of VOCs emitted by Solanum lycopersicum cv Red Setter under water stress conditions was analysed on conspecific plants, which were subsequently subjected to water stress as well, as described in Chapter 1. Subsequently, two different reverse genetics approaches were employed to investigate the involvement of two proteins in VOC perception. The candidates were identified through bioinformatic analysis focusing on their structural potential to bind VOCs. The first experiment focused on plants selected through the TILLING platform, carrying a mutation in the Extra-large guanine nucleotide-binding 3 protein (XLG3) (Solyc08g005310.4.1). This study revealed new potential implications of this important signalling protein in VOC-mediated pathways. Extensive work was then dedicated to the genome editing procedure, applying the CRISPR/Cas9 methodology and Agrobacterium tumefaciens EHA105-mediated transformation in tomato plants (S. lycopersicum cv Red Setter and S. lycopersicum cv Microtom) with the aim of silencing gene encoding for the second protein individuated: RPP13-like protein (Solyc03g078300.1.1). The successful outcome of the genome editing and line propagation, allowed for the execution of a third experiment involving a defRPP13-like line. In this case, the plants were exposed to a single synthetic VOC, the terpenoid β-ionone, and subsequently subjected to biotic stress through inoculation with Botrytis cinerea. Differences in the transcriptome, VOCs emissions, physiological response and damage severity, allowed to highlight the probable involvement of the terpenoid in the regulation of the RPP13-like protein. In all the experiments, were collected data on the effects of exogenous VOCs on the plant transcriptome, through RNA-Seq, on the physiological parameters such as gas exchange and photosynthetic efficiency, using the GFS-3000 system (Walz, Effeltrich, Germany), and on the VOC emissions, using PTR-Qi-TOF MS (Ionicon Analytik GmbH, Innsbruck, Austria were analyzed). For the first two experiments, data from image analysis (High Throughput Plant Phenomics Platform - HTP) were also obtained using the LemnaTec Scanalyzer 3D system, while for the experiment with B. cinerea, direct damage severity was assessed.