Sulfur deficiency-induced genes affect seed protein accumulation and composition under sulfate deprivation.

Sulfur deficiency-induced proteins SDI1 and SDI2 play a fundamental role in sulfur homeostasis under sulfate-deprived conditions (-S) by downregulating glucosinolates. Here, we identified that besides glucosinolate regulation under -S, SDI1 downregulates another sulfur pool, the S-rich 2S seed storage proteins in Arabidopsis (Arabidopsis thaliana) seeds. We identified that MYB28 directly regulates 2S seed storage proteins by binding to the At2S4 promoter. We also showed that SDI1 downregulates 2S seed storage proteins by forming a ternary protein complex with MYB28 and MYC2, another transcription factor involved in the regulation of seed storage proteins. These findings have significant implications for the understanding of plant responses to sulfur deficiency.

The SLIM1 transcription factor affects sugar signaling during sulfur deficiency in Arabidopsis.

The homeostasis of major macronutrient metabolism needs to be tightly regulated, especially when the availability of one or more nutrients fluctuates in the environment. Both sulfur metabolism and glucose signaling are important processes throughout plant growth and development, as well as during stress responses. Still, very little is known about how these processes affect each other, although they are positively connected. Here, we showed in Arabidopsis that the crucial transcription factor of sulfur metabolism, SLIM1, is involved in glucose signaling during shortage of sulfur. The germination rate of the slim1_KO mutant was severely affected by high glucose and osmotic stress. The expression of SLIM1-dependent genes in sulfur deficiency appeared to be additionally induced by a high concentration of either mannitol or glucose, but also by sucrose, which is not only the source of glucose but another signaling molecule. Additionally, SLIM1 affects PAP1 expression during sulfur deficiency by directly binding to its promoter. The lack of PAP1 induction in such conditions leads to much lower anthocyanin production. Taken together, our results indicate that SLIM1 is involved in the glucose response by modulating sulfur metabolism and directly controlling PAP1 expression in Arabidopsis during sulfur deficiency stress.

H+ Transport by K+ EXCHANGE ANTIPORTER3 Promotes Photosynthesis and Growth in Chloroplast ATP Synthase Mutants.

The composition of the thylakoid proton motive force (pmf) is regulated by thylakoid ion transport. Passive ion channels in the thylakoid membrane dissipate the membrane potential (Δψ) component to allow for a higher fraction of pmf stored as a proton concentration gradient (ΔpH). K+/H+ antiport across the thylakoid membrane via K+ EXCHANGE ANTIPORTER3 (KEA3) instead reduces the ΔpH fraction of the pmf. Thereby, KEA3 decreases nonphotochemical quenching (NPQ), thus allowing for higher light use efficiency, which is particularly important during transitions from high to low light. Here, we show that in the background of the Arabidopsis (Arabidopsis thaliana) chloroplast (cp)ATP synthase assembly mutant cgl160, with decreased cpATP synthase activity and increased pmf amplitude, KEA3 plays an important role for photosynthesis and plant growth under steady-state conditions. By comparing cgl160 single with cgl160 kea3 double mutants, we demonstrate that in the cgl160 background loss of KEA3 causes a strong growth penalty. This is due to a reduced photosynthetic capacity of cgl160 kea3 mutants, as these plants have a lower lumenal pH than cgl160 mutants, and thus show substantially increased pH-dependent NPQ and decreased electron transport through the cytochrome b 6 f complex. Overexpression of KEA3 in the cgl160 background reduces pH-dependent NPQ and increases photosystem II efficiency. Taken together, our data provide evidence that under conditions where cpATP synthase activity is low, a KEA3-dependent reduction of ΔpH benefits photosynthesis and growth.

Cysteine and Methionine Biosynthetic Enzymes Have Distinct Effects on Seed Nutritional Quality and on Molecular Phenotypes Associated With Accumulation of a Methionine-Rich Seed Storage Protein in Rice.

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.

CYSTATHIONINE GAMMA-SYNTHASE activity in rice is developmentally regulated and strongly correlated with sulfate.

An important goal of rice cultivar development is improvement of protein quality, especially with respect to essential amino acids such as methionine. With the goal of increasing seed methionine content, we generated Oryza sativa ssp. japonica cv. Taipei 309 transgenic lines expressing a feedback-desensitized CYSTATHIONINE GAMMA-SYNTHASE from Arabidopsis thaliana (AtD-CGS) under the control of the maize ubiquitin promoter. Despite persistently elevated cystathionine gamma-synthase (CGS) activity in the AtD-CGS transgenic lines relative to untransformed Taipei, sulfate was the only sulfur-containing compound found to be elevated throughout vegetative development. Accumulation of methionine and other sulfur-containing metabolites was limited to the leaves of young plants. Sulfate concentration was found to strongly and positively correlate with CGS activity across vegetative development, irrespective of whether the activity was provided by the endogenous rice CGS or by a combination of endogenous and AtD-CGS. Conversely, the concentrations of glutathione, valine, and leucine were clearly negatively correlated with CGS activity in the same tissues. We also observed a strong decrease in CGS activity in both untransformed Taipei and the AtD-CGS transgenic lines as the plants approached heading stage. The mechanism for this downregulation is currently unknown and of potential importance for efforts to increase methionine content in rice.