Gene-trap based identification of stress-activated genes in Arabidopsis thaliana

We employed a large scale gene trap approach to identify genes involved in osmotic stress responses. This strategy allows the analysis of stress-induced Gus expression, so we realized a selective screening of the EXOTIC (EXOn Trapping Insert Consortium) collection, by using Mannitol as selective agent. Moreover this strategy allows the analysis of cell-specific Gus expression, so we searched the EXOTIC lines for guard cell-specific activation of the reporter gene. We identified 3 lines that showed GUS expression only after exposure to Mannitol. In one line we mapped the transposon insertion in the At3g03350 gene, coding for a putative dehydrogenase/reductase protein. At3g03350 is specifically activated by Mannitol in roots, but it is constitutively expressed in the endosperm. We observed an arrest of embryo development at the globular stage in homozygous mutant seeds and verified that it is due to knockout of the At3g03350 gene. Both mutant embryo and endosperm did not show morphological defects. This phenotype suggests that the enzyme is involved in early seed development, particularly in energy metabolism.Subsequently we employed the gene trap approach to identify genes expressed in guard cells. We identified five lines in which the reporter gene was exclusively or preferentially expressed in guard cells. In 3 lines we mapped the DsG insertion in intergenic regions while in 2 lines we mapped the DsG insertion in two annotated genes. CYP86A2, coding for a cytochrome P450 protein, is involved in cutin monomers biosynthesis in stomata and we hypothesize that it is involved in stomatal responses to pathogen attacks. AtPDR3 (pleiotropic drug resistance 3) is an ABC transporter and we provide evidence that it is involved in the modulation of guard cell responses to ABA.