The search for candidate genes associated with natural variation of grain Zn accumulation in barley.

Combating hidden hunger through molecular breeding of nutritionally enriched crops requires a better understanding of micronutrient accumulation. We studied natural variation in grain micronutrient accumulation in barley (Hordeum vulgare L.) and searched for candidate genes by assessing marker-trait associations (MTAs) and by analyzing transcriptional differences between low and high zinc (Zn) accumulating cultivars during grain filling. A collection of 180 barley lines was grown in three different environments. Our results show a pronounced variation in Zn accumulation, which was under strong genotype influence across different environments. Genome-wide association mapping revealed 13 shared MTAs. Across three environments, the most significantly associated marker was on chromosome 2H at 82.8 cM and in close vicinity to two yellow stripe like (YSL) genes. A subset of two pairs of lines with contrasting Zn accumulation was chosen for detailed analysis. Whole ears and flag leaves were analyzed 15 days after pollination to detect transcriptional differences associated with elevated Zn concentrations in the grain. A putative α-amylase/trypsin inhibitor CMb precursor was decidedly higher expressed in high Zn cultivars in whole ears in all comparisons. Additionally, a gene similar to barley metal tolerance protein 5 (MTP5) was found to be a potential candidate gene.

Spatially resolved analysis of variation in barley (Hordeum vulgare) grain micronutrient accumulation.

Genetic biofortification requires knowledge on natural variation and the underlying mechanisms of micronutrient accumulation. We therefore studied diversity in grain micronutrient concentrations and spatial distribution in barley (Hordeum vulgare), a genetically tractable model cereal and an important crop with widespread cultivation. We assembled a diverse collection of barley cultivars and landraces and analysed grain micronutrient profiles in genebank material and after three independent cultivations. Lines with contrasting grain zinc (Zn) accumulation were selected for in-depth analysis of micronutrient distribution within the grain by micro-proton-induced X-ray emission (µ-PIXE). Also, we addressed association with grain cadmium (Cd) accumulation. The analysis of > 120 lines revealed substantial variation, especially in grain Zn concentrations. A large fraction of this variation is due to genetic differences. Grain dissection and µ-PIXE analysis of contrasting lines showed that differences in grain Zn accumulation apply to all parts of the grain including the endosperm. Cd concentrations exceeded the Codex Alimentarius threshold in most of the representative barley lines after cultivation in a Cd-contaminated agricultural soil. Two important conclusions for biofortification are: first, high-Zn grains contain more Zn also in the consumed parts of the grain; and second, higher micronutrient concentrations are strongly associated with higher Cd accumulation.

Silicon-induced reversibility of cadmium toxicity in rice.

Silicon (Si) modulates tolerance to abiotic stresses, but little is known about the reversibility of stress effects by supplementing previously stressed plants with Si. This is surprising since recovery experiments might allow mechanisms of Si-mediated amelioration to be addressed. Rice was exposed to 10 µM CdCl2 for 4 d in hydroponics, followed by 0.6mM Si(OH)4 supplementation for 4 d. Si reversed the effects of Cd, as reflected in plant growth, photosynthesis, elemental composition, and some biochemical parameters. Cd-dependent deregulation of nutrient homeostasis was partially reversed by Si supply. Photosynthetic recovery within 48h following Si supply, coupled with strong stimulation of the ascorbate-glutathione system, indicates efficient activation of defense. The response was further verified by transcript analyses with emphasis on genes encoding members of the stress-associated protein (SAP) family. The transcriptional response to Cd was mostly reversed following Si supply. Reprogramming of the Cd response was obvious for Phytochelatin synthase 1, SAP1 , SAP14, and the transcription factor genes AP2/Erf020, Hsf31, and NAC6 whose transcript levels were strongly activated in roots of Cd-stressed rice, but down-regulated in the presence of Si. These findings, together with changes in biochemical parameters, highlight the significance of Si in growth recovery of Cd-stressed rice and indicate a decisive role for readjusting cell redox homeostasis.

Temporal and Spatial Patterns of Zinc and Iron Accumulation during Barley (Hordeum vulgare L.) Grain Development.

Breeding and engineering of biofortified crops will benefit from a better understanding of bottlenecks controlling micronutrient loading within the seeds. However, few studies have addressed the changes in micronutrient concentrations, localization, and speciation occurring over time. Therefore, we studied spatial patterns of zinc and iron accumulation during grain development in two barley lines with contrasting grain zinc concentrations. Microparticle-induced-X-ray emission and laser ablation-inductively coupled plasma mass spectrometry were used to determine tissue-specific accumulation of zinc, iron, phosphorus, and sulfur. Differences in zinc accumulation between the lines were most evident in the endosperm and aleurone. A gradual decrease in zinc concentrations from the aleurone to the underlying endosperm was observed, while iron and phosphorus concentrations decreased sharply. Iron co-localized with phosphorus in the aleurone, whereas zinc co-localized with sulfur in the sub-aleurone. We hypothesize that differences in grain zinc are largely explained by the endosperm storage capacity. Engineering attempts should be targeted accordingly.