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PLANT PHYSIOLOGY , Vol 103, Issue 2 467-476, Copyright © 1993 by American Society of Plant Biologists

MOLECULAR BIOLOGY AND GENE REGULATION

Cloning of Higher Plant [omega]-3 Fatty Acid Desaturases

N. S. Yadav, A. Wierzbicki, M. Aegerter, C. S. Caster, L. Perez-Grau, A. J. Kinney, W. D. Hitz, J. R. Booth Jr., B. Schweiger, K. L. Stecca, S. M. Allen, M. Blackwell, R. S. Reiter, T. J. Carlson, S. H. Russell, K. A. Feldmann, J. Pierce and J. Browse
Agricultural Products (N.S.Y.,A.W., C.S.C., L.P.-G., A.J.K., J.R.B., B.S., K.L.S., M.B., J.P.) and Central Research and Development (W.D.H., S.M.A., R.S.R., T.J.C., S.H.R., K.A.F.), E. I. duPont de Nemours & Co., Experimental Station, P.O. Box 80402, Wilmington, Delaware 19880-0402

Arabidopsis thaliana T-DNA transformants were screened for mutations affecting seed fatty acid composition. A mutant line was found with reduced 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 cDNA that encodes a polypeptide identified as a microsomal [omega]-3 fatty acid desaturase by its complementation of the mutation. Analysis of lipid content in transgenic tissues demonstrated that this enzyme is limiting 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 desaturation. These Arabidopsis cDNAs were used as hybridization probes to isolate cDNAs encoding homologous proteins from developing seeds of soybean and rapeseed. The high degree of sequence similarity between these sequences suggests that the [omega]-3 desaturases use a common enzyme mechanism.


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