AN EXPRESSION-BASED REVERSE GENETIC STUDY TO IDENTIFY DETERMINANTS OF ABIOTIC STRESS TOLERANCE

Crop productivity is severely affected by drought and high salinity in many regions of the world and the impact of such stresses is predicted to increase drastically in the near future. To guarantee global food production to an increasing world population expected to double by 2050, the identification of genes involved in tolerance mechanisms is a key goal to develop crops better capable to handle these stresses. Adaptation to stress is a complicated process and requires an extensive alteration in gene expression. In a previous study, our group identified several genes whose expression was differentially regulated in Solanum tuberosum culture cells during gradual exposure to increasing concentrations of polyethylene glycol (PEG) (Ambrosone et al., 2013). Among these, we have selected for further studies 50 genes belonging to different functional categories. Orthologous genes in Arabidopsis thaliana were identified based on sequence homology and one homozygous knock-out line was isolated for each gene which contained a T-DNA insertion in the coding sequence. These lines were subjected to a large-scale phenotype screening in order to identify genes with a functional role in adaptation to osmotic stress. Several treatments including NaCl and Abscisic acid (ABA) were conducted and different parameters such as seed germination, root growth and plant survival in the presence of stress evaluated for each genotype. Using this strategy, we have identified 3 genes whose abolished expression has an impact on stress tolerance or ABA sensitivity. In particular, a RNA-binding protein, highly expressed in dry seed and shoot apex, showed tolerance to elevated concentration of ABA. In contrast, survival test and root growth experiments on plates allowed to identify i) a component of the TOR pathway, responsible for activating a cell-growth program in response to nutrients and stresses and ii) a putative subunit of RNA polymerase III which were hyper-sensitive to multiple stresses. We are in the process of performing a detailed functional analysis for the three selected genes using transgenic lines with the purpose of understanding their roles in plant adaptation to osmotic stress and thus contribute to future crop improvement.