CLONING OF HIGHER-PLANT OMEGA-3-FATTY-ACID DESATURASES

Arabidopsis thaliana T-DNA transformants were screened for mutations a ffecting seed fatty acid composition. A mutant line was found with red uced levels of linolenic acid (18:3) due to a T-DNA insertion. Genomic DNA flanking the T-DNA insertion was used to obtain an Arabidopsis cD NA that encodes a polypeptide identified as a microsomal omega-3 fatty acid desaturase by its complementation of the mutation. Analysis of l ipid content in transgenic tissues demonstrated that this enzyme is li miting for 18:3 production in Arabidopsis seeds and carrot hairy roots . This cDNA was used to isolate a related Arabidopsis cDNA, whose mRNA is accumulated to a much higher level in leaf tissue relative to root tissue. This related cDNA encodes a protein that is a homolog of the microsomal desaturase but has an N-terminal extension deduced to be a transit peptide, and its gene maps to a position consistent with that of the Arabidopsis fad D locus, which controls plastid omega-3 desatur ation. These Arabidopsis cDNAs were used as hybridization probes to is olate cDNAs encoding homologous proteins from developing seeds of soyb ean and rapeseed. The high degree of sequence similarity between these sequences suggests that the omega-3 desaturases use a common enzyme m echanism.