2013 Annual Report
1a.Objectives (from AD-416):
Objective 1. Use genetic, molecular, and physiological approaches to define the role of specific genes and gene products in the acquisition and whole-organism partitioning of minerals (iron, zinc, calcium, and magnesium) and other factors that inhibit or promote absorption of these minerals in plant foods.
Sub-objective 1.A. Identify quantitative trait loci (QTLs) associated with elevated seed or leaf mineral concentrations or seed biomass accumulation.
Sub-objective 1.B. Identify rate-limiting and/or novel genes that contribute to tissue Fe, Zn, Ca, and Mg concentrations or seed biomass accumulation.
Sub-objective 1.C. Assess the functional role of newly identified gene products in whole-plant nutrition and nutrient partitioning.
Sub-objective 1.D. Measure the effects of modulating cation transporter functions on plant Ca, Mg, Fe and Zn content.
Sub-objective 1.E. Measure the impact of altered transport function in agriculturally important crops.
Sub-objective 1.F. Identify and isolate genes that are involved in calcium oxalate crystal formation in selected mutants.
Sub-objective 1.G. Assess the role of the identified genes in calcium oxalate crystal formation.
Objective 2. Conduct animal and human feeding studies to determine mineral bioavailability of the nutritionally enhanced crops.
Sub-objective 2.A. Use novel transgenic plants to dissect the relationship between mineral partitioning and nutrient absorption in mice feeding studies.
Sub-objective 2.B. If the rodent studies demonstrate proof of concept, initiate pilot feeding studies using young adults.
Objective 3. Develop new, cost-effective methods for the intrinsic labeling of plant foods for use in nutrient bioavailability studies.
1b.Approach (from AD-416):
The long-term objective of this project is to contribute to the development of nutritionally enhanced plant foods and to develop tools for testing nutrient bioavailability. We have chosen to work initially with plants that are tractable molecular genetic systems (Arabidopsis, Medicago, and soybean) where we can perform gene discovery quickly. We then translate these findings into agriculturally important crops that can be easily transformed and for which established protocols are in place for measuring nutrient absorption in both mice and humans. Specifically, we will work to identify and characterize genes and gene products that are involved in mineral transport throughout the plant, focusing both on whole organ accumulation and subcellular partitioning of minerals. We also will identify and characterize the molecular processes associated with calcium oxalate formation in plants. We envision this work to eventually have relevance to mineral nutrition improvement (e.g., calcium, magnesium, iron, and zinc) in several agronomic crops. In addition, we will develop new, cost-effective methods for the intrinsic labeling of plant foods, using stable isotopes, in order to facilitate nutrient bioavailability studies in humans. These efforts will expand our capabilities for assessing the absorption and metabolism of various plant-derived minerals and phytochemicals. They also will facilitate the generation of new bioavailability data for various nutrients, which will allow informed decisions when policymakers establish future dietary recommendations for humans.
Dry beans are an important food source that can provide protein, energy in the form of starch, and several human-essential minerals, including iron. Because of the importance of iron in human health, our goal has been to find ways to increase the concentration of iron in plant food crops such as dry bean, to ensure that consumers meet their daily intake requirements for this mineral. In one set of studies, CNRC researchers investigated the genetic factors that control the distribution of iron in bean seeds. Previous work has shown that iron concentration is higher in the dry bean's seed coat, relative to the cotyledon and embryo axis tissues within that seed coat. We measured mineral concentrations in seed coats of individual bean types that were part of a unique population of bean plants. This population was derived from a wild bean and a cultivated bean. A broad range of mineral concentrations was found in the seed coats of the plants from this population. We were able to use these results to identify regions of the bean plant's chromosomes (i.e., DNA segments) that were associated with elevated iron concentrations in seed coats. This information will help us identify genes that control iron distribution and concentrations in seeds. It will also provide tools for breeders to develop nutritionally improved cultivars of dry beans.
Corn is an important grain crop that provides energy, protein, minerals, and vitamins for humans. Grain quality, however, can be affected by growing conditions, such as soil fertility and water availability. Because we would like to develop new, improved corn varieties of the highest potential nutritional value, we were interested in understanding how different environmental conditions might affect grain mineral concentrations, and iron concentration in particular. We grew several diverse corn varieties in field plots that were provided with full nitrogen nutrition and ample water, or were limited in nitrogen, water, or both. We found that the corn varieties with the highest iron concentration differed, depending on the environmental conditions they were grown under. This work tells us that breeding for improved grain nutritional quality will require testing under a range of field conditions and environments. Our results are promising, because they will help us understand which environments are most important to make meaningful nutritional improvements in future corn varieties.
Potato is a food crop that can provide minerals in addition to energy (as starch) and some protein. An ARS collaborator in Prosser, WA, had identified different potato lines that had high or low levels of iron in their tubers. Our lab has studied root processes that help to bring iron into the plant, to identify the reason for the differences in iron concentrations in the tubers. Specifically, we have studied a root process that converts soil iron into a form that can be absorbed by the potato plant. High iron potatoes were found to have a higher capacity for this process. We are now trying to find a gene in potato that is linked to this process. This understanding will provide us with new tools to develop nutritionally improved cultivars of potato.
The proliferation of DNA sequence information has provided new tools that can be used to identify genes associated with useful traits, especially when these tools are combined with the study of diverse cultivars of a given species. In one study, we used a large set of diverse rice lines from the USDA Rice Minicore Collection to measure seed mineral concentrations and seed growth and mineral import, to aid us in identifying regions of the plant's genome that were associated with grain mineral density or seed growth. Several regions along the rice plant's chromosomes were identified to be associated with seed growth traits or enhanced seed mineral accumulation. These results should provide new genes and new breeding tools to help scientists improve the nutritional quality of rice. Enhancing the nutritional quality of crops is of US and international importance, and multiple methods have been utilized to increase the nutrient content of legume seeds. Because the movement of nutrients from leaves to growing reproductive tissues greatly contributes to the final composition of the mature seeds, manipulating the movement of minerals stored in leaves is a potential strategy for increasing final seed mineral concentration. It has previously been reported that seeds of the model legume, Medicago truncatula (type A17), contain a lower iron concentration compared to another type (DZA315.16). We hypothesized that this difference was due to variation in leaf processes and sought to understand these distinctions by challenging both plant types with long-term iron deficiency. As expected, when challenged with iron deficiency for a month, growth of both plant types significantly decreased relative to controls; however, there were differences between the plant types in the number of leaves retained on the plant and the number of pods and seeds that were successfully developed. We also studied the expression of various iron nutrition related genes in leaves of the two plant types. These results will help us identify how plants cope with limited iron availability and the impact of this stress on seed yield and seed nutritional quality.
Many edible plants form crystals of a calcium oxalate. The calcium present in these crystals is unavailable for nutritional absorption by humans and other animals. One strategy to improve the nutritional value of edible plants, in terms of calcium availability, is to reduce the amount of calcium bound within these oxalate crystals. Before such a strategy can be designed, a better understanding of the molecular mechanisms regulating calcium oxalate crystal formation is required. As a step toward gaining such an understanding, we have completed genetic and molecular characterizations of a newly identified gene required for crystal accumulation in the model legume Medicago truncatula. We utilized a small DNA fragment of a mutant gene to isolate the coding sequence for this gene's transcript and conducted gene expression analysis. We have completed the generation of necessary molecular tools to down-regulate the expression of this gene and have initiated efforts to use this approach with wild-type Medicago truncatula plants. We anticipate that these studies will help lead to the development of a strategy to reduce the amount of calcium bound in the oxalate crystal and improve calcium bioavailability in other edible legumes.
The influence of the distribution of plant nutrients on their bioavailability and therefore on human health is poorly understood. Previously we have demonstrated that plants that have defects in packaging calcium into crystal forms are actually healthier than the same plants that make these crystals. CNRC researchers conducted long-term feeding studies using these modified plants fed to animals with defects in bone health and calcium metabolism. We show that diets containing these plants significantly improve bone health and calcium metabolism in these animals. Our study suggests that plant-based diets lacking calcium packaging mechanisms may be used as a preventative or corrective treatment to suppress disorders associated with improper calcium metabolism.
Location, Location, Location - where a protein resides within a plant cell. Determining where a given protein is located within a cell often provides clues about its function. To help determine where a given protein is located within a cell, Children's Nutrition Research Center researchers in Houston, Texas, have generated a set of green fluorescence protein (GFP) marker lines in the plant Medicago truncatula that allows visualization of the different parts of the plant cell. It is expected that this marker set will prove to be a useful resource in the study of any given biological pathway, and thus a germplasm release has been completed for the entire marker set so it can be available for use to the plant research community. Currently, we are utilizing this marker set to uncover the function of proteins that are required for calcium oxalate crystal formation. The calcium bound in the crystals of calcium oxalate has been shown to be unavailable for nutritional absorption by humans and other animals. By gaining a better understanding of how plants form these crystals we can design strategies to decrease the amount of calcium bound in these crystals and improve the nutritional quality of plant foods.
Calcium fertilizer is important for seed nutritional quality in peanut. Peanuts are grown in sandy soils, which often have low levels of plant-required mineral nutrients such as calcium. For many soil types, peanut farmers apply a calcium fertilizer (gypsum) to improve peanut yields and the quality of the harvested seeds; however, there is little information on the effect of soil calcium application on the plant's uptake of other minerals, or the delivery of these minerals to the pod walls and seeds. Using different peanut varieties and different calcium fertilizer treatments, plant researchers at the Children's Nutrition Research Center in Houston, Texas, found that calcium applications resulted in higher concentrations of calcium, sulfur, and zinc in seeds, but showed slight decreases in phosphorus and sodium concentrations. There were no effects on any other minerals studied. These results provide useful information on the growing conditions and agricultural methods needed to maintain peanut yield and seed nutritional quality in the future.