Abstract
Staple crops in human and livestock diets suffer from deficiencies in certain “essential” amino acids including methionine. With the goal of increasing methionine in rice seed, we generated a pair of “Push × Pull” double transgenic lines, each containing a methionine-dense seed storage protein (2S albumin from sunflower, HaSSA) and an exogenous enzyme for either methionine (feedback desensitized cystathionine gamma synthase from Arabidopsis, AtD-CGS) or cysteine (serine acetyltransferase from E. coli, EcSAT) biosynthesis. In both double transgenic lines, the total seed methionine content was approximately 50% higher than in their untransformed parental line, Oryza sativa ssp. japonica cv. Taipei 309. HaSSA-containing rice seeds were reported to display an altered seed protein profile, speculatively due to insufficient sulfur amino acid content. However, here we present data suggesting that this may result from an overloaded protein folding machinery in the endoplasmic reticulum rather than primarily from redistribution of limited methionine from endogenous seed proteins to HaSSA. We hypothesize that HaSSA-associated endoplasmic reticulum stress results in redox perturbations that negatively impact sulfate reduction to cysteine, and we speculate that this is mitigated by EcSAT-associated increased sulfur import into the seed, which facilitates additional synthesis of cysteine and glutathione. The data presented here reveal challenges associated with increasing the methionine content in rice seed, including what may be relatively low protein folding capacity in the endoplasmic reticulum and an insufficient pool of sulfate available for additional cysteine and methionine synthesis. We propose that future approaches to further improve the methionine content in rice should focus on increasing seed sulfur loading and avoiding the accumulation of unfolded proteins in the endoplasmic reticulum.Oryza sativa ssp. japonica: urn:lsid:ipni.org:names:60471378-2.
Highlights
Unlike plants, all animals lack the enzymatic machinery to synthesize de novo some of the 20 proteinaceous amino acids
The AtD-CGS and EcSAT transgenes are both driven by a maize ubiquitin promoter, and their transcripts were highly abundant in milkripe seeds, even after at least five generations post-transformation (Figures 1A, B)
We generated so-called “Push × Pull” lines by combining a strong sulfur amino acids (SAA) sink with cysteine or methionine biosynthesis transgenes with the idea that increased production of S-AA would allow for high levels of HaSSA accumulation without downregulation of endogenous relatively S-rich seed storage proteins (SSP)
Summary
All animals lack the enzymatic machinery to synthesize de novo some of the 20 proteinaceous amino acids. These so called “essential amino acids” must be consumed in their diet. For optimal growth, these essential amino acids must be consumed in the right balance for the animal’s metabolic needs. Methionine (Met) is one such essential amino acid, but the amount present in plant-based animal feed blends is typically insufficient optimal livestock growth and health. Increasing the methionine and cysteine content of commodity cereals and grain legumes would benefit farmers by elevating the value of their crop and would be of benefit to livestock rearing by reducing the need for synthetic amino acid supplementation of animal feed
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