A transgene for high methionine protein is posttranscriptionally regulated by methionine

beta-Zein is one of the seed storage proteins of maize that is high in methionine (Met). In alfalfa, the beta-zein gene driven by the CaMV 35S promoter showed an 8-fold lower level of transcript and protein when compared with the level in tobacco transformed with the same gene construct. The reporter gene (GUS) driven by the CaMV 35S promoter showed only a 4-fold difference between alfalfa and tobacco, suggesting that the expression of the beta-zein gene is posttranscriptionally regulated in alfalfa. Callus of alfalfa transformants with the beta-zein gene construct treated with exogenous Met, showed a significant increase in the beta-zein level, suggesting that free Met may be limiting in the synthesis of beta-zein in alfalfa. The introduction of the Arabidopsis thaliana cystathionine gamma-synthase (AtC gamma S) gene driven by the CaMV 35S promoter into alfalfa showed a significant increase in the level of free Met and its metabolite, S-methyl methionine (SMM), but not in the bound fraction. Coexpression of AtC gamma S and beta-zein in alfalfa increased the level of beta-zein transcript and protein and decreased free Met, which suggests that the beta-zein is posttranscriptionally regulated by free Met. The expression of AtC gamma S in tobacco did not produce a significant increase in free Met or SMM and coexpression of AtC gamma S and beta-zein did not result in changes in the beta-zein level. The results demonstrate the efficacy of the synergistic approach of increasing both the sink and the source for increasing the levels of high Met beta-zein.