Cloning and molecular characterization of the delta6-desaturase from two echium plant species: production of GLA by heterologous expression in yeast and tobacco.

Abstract

The synthesis of GLA (delta6,9,12-1-8:3) is carried out in a number of plant taxa by introducing a double bond at the delta6 position of its precursor, linoleic acid (delta9,12-18:2), through a reaction catalyzed by a delta6-desaturase enzyme. We have cloned genes encoding the delta6-desaturase (D6DES) from two different Macaronesian Echium species, E. pitardii and E. gentianoides (Boraginaceae), which are characterized by the accumulation of high amounts of GLA in their seeds. The Echium D6DES genes encode proteins of 438 amino acids bearing the prototypical cytochrome b(5) domain at the N-terminus. Cladistic analysis of desaturases from higher plants groups the Echium D6DES proteins together with other delta6-desaturases in a different cluster from that of the highly related delta8-desaturases. Expression analysis carried out in E. pitardii shows a positive correlation between the D6DES transcript level and GLA accumulation in different tissues of the plant. Although a ubiquitous expression in all organs is observed, the transcript is particularly abundant in developing fruits, whereas a much lower level is present in mature leaves. Functional characterization of the D6DES gene from E. gentianoides has been achieved by heterologous expression in tobacco plants and in the yeast Saccharomyces cerevisiae. In both cases, overexpression of the gene led to the synthesis of GLA. Biotechnological application of these results can be envisaged as an initial step toward the generation of transgenic oleaginous plants producing GLA.