Genetic variability in the promoter of miR397 in Picea abies

Norway spruce (Picea abies Karst.) is a tree species that belongs to conifers, a taxon that is extremely important both from an ecological and an economical point of view. Being perennial, this species has often to face suboptimal environmental conditions and to adapt to them: microRNAs are a fundamental class of regulatory molecules, often involved in stress responses and therefore, potentially very important for plant adaptive processes. The focus of this study is on miR397: in Arabidopsis thaliana miR397 was shown to be involved in the regulation of copper homeostasis and of the transcription of laccases. These enzymes operate during lignin biosynthesis and therefore their regulation is really important in woody plants like Norway spruce, in order to react to mechanical stress and to resist to the attack of pathogens. In order to understand miR397 regulatory mechanisms, its promoter was isolated in this species and putative regulatory elements were identified. This region, together with the microRNA stem-loop region, was sequenced in seeds produced by individuals originating from different alpine populations in Italy, Austria and Switzerland. In the mature miR397, the most important part for microRNA regulatory function, no polymorphism was found in the analyzed samples. This result suggests that purifying selection is probably acting on this sequence in order to preserve microRNA functionality. As regards the promoter region, several single nucleotide polymorphisms (SNPs) and some insertions/deletions were identified. Some of them are located in the putative regulatory elements, therefore they are good candidates to test if they influence the microRNA expression level and if they have consequences on phenotype that can be relevant in the process of adaptation. This will provide deeper insights into the adaptive role of microRNAs in Conifers