Abstract
Expression of heterologous methionine-rich proteins to increase the overall sulfur amino acid content of soybean seeds has been only marginally successful, presumably due to low accumulation of transgenes in soybeans or due to gene silencing. Proteome rebalancing of seed proteins has been shown to promote the accumulation of foreign proteins. In this study, we have utilized RNAi technology to suppress the expression of the β-conglycinin, the abundant 7S seed storage proteins of soybean. Western blot and 2D-gel analysis revealed that β-conglycinin knockdown line (SAM) failed to accumulate the α′, α, and β-subunits of β-conglycinin. The proteome rebalanced SAM retained the overall protein and oil content similar to that of wild-type soybean. We also generated transgenic soybean lines expressing methionine-rich 11 kDa δ-zein under the control of either the glycinin or β-conglycinin promoter. The introgression of the 11 kDa δ-zein into β-conglycinin knockdown line did not enhance the accumulation of the 11 kDa δ-zein. However, when the same plants were grown in sulfur-rich medium, we observed 3- to 16-fold increased accumulation of the 11 kDa δ-zein. Transmission electron microscopy observation revealed that seeds grown in sulfur-rich medium contained numerous endoplasmic reticulum derived protein bodies. Our findings suggest that sulfur availability, not proteome rebalancing, is needed for high-level accumulation of heterologous methionine-rich proteins in soybean seeds.
Highlights
Due to their high protein content and relatively low cost, soybeans (Glycine max (L.) Merr.) are used as an animal feed throughout the world
Our results indicate that the availability of sulfur, not proteome rebalancing, is more critical for high-level accumulation of methionine-rich proteins in soybean seeds
CREATION OF B-CONGLYCININ KNOCKDOWN SOYBEAN SEEDS The suppression of the 7S globulins accumulation in soybean seeds was accomplished by RNAi utilizing Agrobacteriummediated cotyledonary node transformation protocol (Hinchee et al, 1988)
Summary
Due to their high protein content and relatively low cost, soybeans (Glycine max (L.) Merr.) are used as an animal feed throughout the world. The abundant storage proteins of soybean are the salt-soluble globulins, the 7S β-conglycinin and the 11S glycinin, which together could account for about 70% of the total seed proteins (Nielsen, 1996; Nielsen and Nam, 1999; Krishnan, 2000). Both these classes of proteins are encoded by multiple gene families (Harada et al, 1989; Nielsen et al, 1989). The β-conglycinins are glycoproteins (Thanh and Shibasaki, 1978) Because of their abundance, the 7S and
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