Abstract
Folates are indispensable co-factors for one-carbon metabolism in all organisms. In humans, suboptimal folate intake results in serious disorders. One promising strategy for improving human folate status is to enhance folate levels in food crops by metabolic engineering. In this study, we cloned two GmGCHI (GTP cyclohydrolase I) genes (Gm8gGCHI and Gm3gGCHI) and one GmADCS (aminodeoxychorismate synthase) gene from soybean, which are responsible for synthesizing the folate precursors pterin and p-aminobenzoate, respectively. We initially confirmed their functions in transgenic Arabidopsis plants and found that Gm8gGCHI increased pterin and folate production more than Gm3gGCHI did. We then co-expressed Gm8gGCHI and GmADCS driven by endosperm-specific promoters in maize and wheat, two major staple crops, to boost their folate metabolic flux. A 4.2-fold and 2.3-fold increase in folate levels were observed in transgenic maize and wheat grains, respectively. To optimize wheat folate enhancement, codon-optimized Gm8gGCHI and tomato LeADCS genes under the control of a wheat endosperm-specific glutenin promoter (1Dx5) were co-transformed. This yielded a 5.6-fold increase in folate in transgenic wheat grains (Gm8gGCHI+/LeADCS+). This two-gene co-expression strategy therefore has the potential to greatly enhance folate levels in maize and wheat, thus improving their nutritional value.
Highlights
Tetrahydrofolate and its derivatives are collectively termed folates and they belong to a group of water-soluble vitamin B compounds (B9) (Hanson and Gregory, 2002; Basset et al, 2005)
By using the Arabidopsis AtGCHI and AtADCS coding sequences as query sequences, we found several short and disconnected expressed sequence tag (EST) related to soybean GTP cyclohydrolase I (GCHI) and aminodeoxychorismate synthase (ADCS) (Supplementary Fig. S1)
The two GmGCHI genes were located on chromosome 3 (3g21540) and chromosome 8 (8g46160), and were named Gm3gGCHI and Gm8gGCHI, respectively, and GmADCS was located on chromosome 10 (10g35580)
Summary
Tetrahydrofolate and its derivatives are collectively termed folates and they belong to a group of water-soluble vitamin B compounds (B9) (Hanson and Gregory, 2002; Basset et al, 2005). Folates play roles in histidine degradation pathways in mammals, including humans (Hilton et al, 2003). Folates are necessary for biosynthesis of lignin, alkaloids, and chlorophyll (Hanson and Gregory, 2011).The significant roles played by folates indicate that they are indispensable micronutrients for all living organisms. Microbes and plants can synthesize folates de novo, but mammals (including humans) lack the complete biosynthesis system and are unable to produce folates. They must rely on dietary sources, mainly from plants. Liver, green leafy vegetables, and leguminous vegetables are high in folates, whereas some staple
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