Soybean seeds expressing feedback-insensitive cystathionine γ-synthase exhibit a higher content of methionine

Abstract

Soybean seeds provide an excellent source of protein for human and livestock nutrition. However, their nutritional quality is hampered by a low concentration of the essential sulfur amino acid, methionine (Met). In order to study factors that regulate Met synthesis in soybean seeds, this study used the Met-insensitive form of Arabidopsis cystathionine γ-synthase (AtD-CGS), which is the first committed enzyme of Met biosynthesis. This gene was expressed under the control of a seed-specific promoter, legumin B4, and used to transform the soybean cultivar Zigongdongdou (ZD). In three transgenic lines that exhibited the highest expression level of AtD-CGS, the level of soluble Met increased significantly in developing green seeds (3.8-7-fold). These seeds also showed high levels of other amino acids. This phenomenon was more prominent in two transgenic lines, ZD24 and ZD91. The total Met content, which including Met incorporated into proteins, significantly increased in the mature dry seeds of these two transgenic lines by 1.8- and 2.3-fold, respectively. This elevation was accompanied by a higher content of other protein-incorporated amino acids, which led to significantly higher total protein content in the seeds of these two lines. However, in a third transgenic line, ZD01, the level of total Met and the level of other amino acids did not increase significantly in the mature dry seeds. This line also showed no significant change in protein levels. This suggests a positive connection between high Met content and the synthesis of other amino acids that enable the synthesis of more seed proteins.

Seed-Specific Expression of a Feedback-Insensitive Form of CYSTATHIONINE-γ-SYNTHASE in Arabidopsis Stimulates Metabolic and Transcriptomic Responses Associated with Desiccation Stress

“…The higher levels of soluble amino acids triggered their incorporation into seed storage proteins, leading to the elevation of total protein contents. Similar results were also found in two lines of transgenic soybean (Glycine max) and tobacco seeds expressing the AtD-CGS Song et al, 2013). However, such elevations were not observed in transgenic seeds with elevated levels of other amino acids.…”

“…The results of this study show that higher expression levels of AtD-CGS elevate the levels of soluble and total Met, indicating that Arabidopsis seeds, like legume and tobacco seeds (Hanafy et al, 2013;Matityahu et al, 2013;Song et al, 2013), are able to synthesize Met de novo through the Asp family pathway. However, further investigation is required to reveal the relative contribution of the two pathways of Met synthesis (Asp and SMM) in Arabidopsis seeds, since significant increases in the expression of HMT1 and HMT3 in the SMM cycle were detected.…”

“…For total protein and lipid determinations, 100 mg of mature dry seeds was collected from four distinct biological pools and treated according to the Kjeldahl and Soxhlet methods, as described previously (Song et al, 2013). The same amounts were used to calorimetrically measure total reducing sugars after carbohydrate and starch hydrolysis using the method of Sumner (1924).…”

“…1). Recent studies performed in legumes and tobacco (Nicotiana tabacum) seeds show that seed-specific expression of feedback-insensitive mutated forms of Arabidopsis (Arabidopsis thaliana) CGS (AtCGS) leads to significantly higher levels of soluble Met (Hanafy et al, 2013;Matityahu et al, 2013;Song et al, 2013), indicating that Met can be synthesized de novo in these seeds via the Asp family pathway through CGS activity in a similar manner to that in vegetative tissues. However, studies performed on Arabidopsis and wheat (Triticum aestivum) seeds suggest the existence of an alternative pathway of Met synthesis in seeds, in which Met synthesized in vegetative tissues is converted to SMM, which is then transported through the phloem into the developing seeds and reconverted back to soluble Met by the activity of HMT (Bourgis et al, 1999;Lee et al, 2008;Fig.…”

High-methionine soybean has no significant effect on nitrogen-transforming bacteria in rhizosphere soil

“…ZD91 contains the Arabidopsis cystathionine γ-synthase gene, introduced artificially into the soybean cv. Zigongdongdou, and produces a high methionine content in seeds (Song et al 2013).…”