Sequence characterization and temporal expression analysis of different SADs and FAD2-2 genes in two Iranian olive cultivars

Abstract

Stearoyl-acyl carrier protein (ACP) desaturases (SADs) and fatty acid desaturases (FADs) are the main genes catalyzing the first and the second desaturation steps throughout the biosynthesis of oleic and linoleic acids, respectively, hence, the major determinants of oil quality and composition. To uncover the molecular mechanisms behind differential oil content and composition in olive fruit, the nucleotide and amino acid sequences of FAD2-2 and three SAD genes were characterized in 'Mari' and 'Shengeh' as two Iranian olive cultivars displaying high contrasting oil composition. In addition, the expression levels of these genes were screened at different time points during fruit development and ripening. Despite detection of a number of nucleotide sequence variations in four characterized genes, the results of amino acid analyses have shown most of them were synonymous and caused no differences at protein level. However, some of the single nucleotide variations caused non cognate amino acid substitution indicating possible conformational changes in the resultant peptide. In particular, regarding OeFAD2-2, a nonsynonymous SNP was detected in 'Shengeh' compared with other olive's FAD2-2 causing an amino acid lysine residue substitution for a glutamic acid, as well as a nonsynonymous substitution in 'Mari' (threonine to serine). The in silico prediction of three-dimensional structure of FAD2-2 revealed these substitutions may lead to structural changes in the final protein structure and function, hence, may contribute to the higher activity of FAD2-2 protein in mesocarp of 'Shengeh' than 'Mari' cultivars. According to results obtained from sequence analysis of oil biosynthesis genes, two Iranian olive cultivars were the most similar and had high sequence differences with international varieties. Results of temporal transcript abundance revealed that OeSAD2 had higher expression in 'Mari' (the high quality oil cultivar) than 'Shengeh' during the critical period of oil biosynthesis. Results of our data corroborates the previous records suggesting that OeSAD2 as the most pronounced isoform of stearoyl-acyl carrier protein desaturases responsible for oleic acid biosynthesis in olive mesocarp. In addition our results also suggest that OeSAD3 may have contributed to the higher oil quality of 'Mari' than 'Shengeh'. Based on our results, OeSAD2 and OeFAD2-2 are suggested as the possible targets for engineering fatty acid composition to help pyramid genes to improve quality and quantity of olive oil and creating high quality olive cultivars.