Poster: Metabolic Engineering

Abs # 1016: Sulfate Reduction Limits Methionine and Cysteine Content in Maize Seed

Presenter:	Tarczynski, Mitchell C, mitchell.tarczynski@pioneer.com
Authors	Tarczynski, Mitchell C (A)   Martin, Melinda  (B)   Shen, Bo  (A)   Li, Changjiang  (A)   Leustek, Thomas  (B)   Falco, Carl  (C)   Glassman, Kimberly  (A)   Ranch, Jerry  (A)   Allen, William  (A)
Affiliations:	(A): Pioneer Hi-Bred Inc., Intl. a DuPont Co, Johnston, IA (B): Rutgers University, New Brunswick, NJ (C): DuPont Company, Wilmington, DE

The sulfur-containing amino-acids methionine (Met) and cysteine (Cys) are required in animal diets. Increasing the accumulation of these amino acids in maize grain would increase the nutritional quality of this staple. Significant increases in seed Met content have been obtained through the expression of storage proteins rich in Met. However, in at least soybean and maize, much of the increased Met in seed can be accounted for by a loss of Cys, suggesting that Cys production/availability quickly becomes limiting for Met synthesis. Studies of the sulfur status of maize seed indicated that the sulfate reduction pathway was a major limitation in sulfur amino-acid biosynthesis. To test this possibility, we constitutively expressed and targeted to the chloroplast two bacterial adenylylsulfate reductases (5’-adenylylsulphate reductase from P. aeruginosa and 3’-phosphoadenylylsulfate reductase from E. coli) in maize, in an effort to deregulate sulfite formation. Vegetative portions of transgenic plants accumulated inorganic forms of sulfur and reduced thiol compounds; both enzymes were capable of causing such changes. Free Met and Cys did not accumulate substantially in vegetative tissues or seed; however, the level of total seed Met and Cys increased by 200% and 140%, respectively. The increased Met and Cys in maize seed could be attributed to elevated expression and accumulation of the endosperm-localized, sulfur-rich zeins. Reciprocal crossing experiments indicated that the increase in total Met and Cys was the result of increased sulfur amino-acid biosynthesis in vegetative maternal tissues rather than in endosperm. Experiments with additional genes that appear to have a role in limiting sulfur amino-acid biosynthesis will also be presented.