2012 Annual Report
1a.Objectives (from AD-416):
1) Identify genes and associated physiological mechanisms for aluminum tolerance in the important cereal crop species, maize and sorghum, with the long-term goal of improving crop production on acid soils..
2)Describe molecular and physiological mechanisms of heavy metal/micronutrient tolerance and transport in the metal hyperaccumulator, Thlaspi caerulescens, and evaluate how these gene systems can be used for phytoremediation of metal-contaminated soils and for enhancing micronutrient nutrition of food crops.
1b.Approach (from AD-416):
1) Sorghum represents plant species where Al tolerance is a simple trait. We have recently cloned the major sorghum Al tolerance gene, AltSB, and found it is a novel solute transporter. The function of AltSB will be studied using a multifaceted approach including the effect of increased/decreased AltSB expression on the physiology of Al tolerance, association analysis correlating sequence and phenotypic variation of multiple AltSB alleles, and analysis of AltSB transporter properties when expressed in heterologous systems..
2)Maize represents a plant species where Al tolerance is a complex, quantitative trait. We have identified a number of Al tolerance QTL in maize, and will work towards cloning these QTL via a combination of gene and protein expression analysis, high resolution mapping, and analysis of candidate tolerance genes based on homology to Al tolerance genes recently cloned in sorghum and wheat..
3)An investigation of the role of hyperexpression of a suite of micronutrient and heavy metal-related genes in heavy metal hyperaccumulation in Thlaspi caerulescens will involve investigation of cis and trans factors that control micronutrient (Zn) homeostasis in the related non-accumulator, Arabidopsis thaliana, and how these elements are altered in T. caerulescens to contribute to the enhanced metal accumulation and tolerance..
4)We have recently identified several genes that play important roles in the hyperaccumulation phenotype in T. caerulescens, including a heavy metal ATPase and a protein kinase, and the functioning of these genes in heavy metal hyperaccumulation, as well as in micronutrient nutrition will be studied.
Under sub-objective 1A, which deals with maize aluminum (Al) tolerance, ARS researchers at the Robert W. Holley Center for Agriculture & Health in Ithaca, NY, studied the regulation of ZmMATE1 expression, the first identified maize Al tolerance gene which is closely related to our sorghum Al tolerance gene, SbMATE. Like SbMATE, it is a root transporter which mediates citric acid release from the root tip into the soil where the citric acid binds and detoxifies Al ions in the soil, allowing the root to grow. It was discovered that the high ZmMATE1 expression in root tips of Al tolerant maize lines is due to the presence of 3 nearly identical copies of ZmMATE1, and all three copies are functional. Al sensitive maize lines only have 1 copy of the ZmMATE1 gene in the root tip.
For sub-objective 1B, we identified a novel mechanism of regulation of the sorghum Al tolerance protein, SbMATE. We identified a second protein, SbMBP (SbMATE binding protein), that binds very strongly to the SbMATE protein and regulates its function. We found that the binding of SbMBP to SbMATE blocks citrate transport and that SbMBP is an Al sensor and when it binds Al ions, it no longer binds to SbMATE, allowing the transport of citrate out of the root. This regulation of SbMATE ensures that unnecessary carbon loss from the root does not occur under non-Al toxic conditions, as citrate release from the root is a significant cost to the plant. In collaboration with Embrapa Maize and Sorghum, Brazil, we are using these findings to improve productivity of sorghum grown as a major food crop in sub-Saharan Africa and investigating the possibility to increase the productivity of biofuel sorghum grown on acid soils prevalent in the southeastern U.S.
For sub-objective 1C, we are improving our techniques for the imaging and reconstruction of root system architecture (RSA), which is important for acquisition of limiting nutrients like N, P and water, in 3 dimensions. Last year, our system to grow cereal roots in transparent gellan gum tubes was described that allows us to digitally image RSA in great detail and use our RootReader 3D software system to reconstruct the multiple images of the roots into a three dimensional model of the whole root system. The system was used successfully in the genetic mapping of rice RSA traits. For sorghum, we have developed a new and improved 3D growth system. The seedlings are now grown in glass cylinders in nutrient solution and not gellan gum. The 3D root system architecture is maintained in this hydroponic system by the series of plastic grids spaced at vertical intervals in the glass growth cylinder, allowing the roots to grow freely but as they grow through the mesh, the 3D architecture is maintained as the root systems are digitally imaged. This approach opens up many more avenues for our RSA research. For example, now we can readily modify the nutrient solution imposing P or N deficiency, or osmotic stress. Also, we can now image root systems of significantly older plants which allows us, for example, to study later developing root types such as sorghum crown roots that are important for water acquisition.
A second novel sorghum protein also improves sorghum yields on acid soils. Acid soils make up as much as 50% of the world’s soils, particularly in the tropics and subtropics where many developing countries are located. Also significant areas in the eastern and southern US have highly acidic soils. ARS researchers at the Robert W. Holley Center for Agriculture and Health at Ithaca, New York, have discovered a second novel protein in sorghum that works in tandem with another sorghum protein they discovered allows sorghum to grow on acid soils. The discovery of these two proteins can be used to more effectively develop high yielding sorghum and other cereal crops to be grown on acid soils in developing countries including sub-Saharan Africa, Southeast Asia and South America, as well as on acid soils in the United States.
New methods developed to identify an important modification to plant proteins. Proteins that control many processes in all organisms undergo specific modifications that are important for their function. One of these modifications is called phosphorylation, where a second protein inserts a phosphate group onto the protein to be regulated. Phosphorylation is critical for the regulation on many proteins in all organisms, including important plant crop species. ARS researchers at the Robert W. Holley Center for Agriculture and Health at Ithaca, New York, have developed new laboratory methods that will allow researchers to rapidly determine if a protein has been phosphorylated and where in the protein this has occurred. This will help us understand how plant proteins that are involved in important crop traits, such as the ability to grow on acid soils, are regulated which in turn may help researchers enhance plant traits regulated by this phosphorylation process.
New methods developed to identify a second important modification to plant proteins. A second modification of plant proteins that is involved in the regulation of protein function is a process where sugar molecules are added to the protein, which is called glycosylation. ARS researchers at the Robert W. Holley Center for Agriculture and Health at Ithaca, New York, have developed new laboratory methods that will allow researchers to rapidly determine if a protein has been glycosylated and to quantify the amount of these modified proteins in important plant crop species. This new technique will enable researchers to better understand the role of this protein modification process in important plant traits and possibly can be used to improve specific traits in plant species.
Improved yields from sorghum grown on high acid soils. Acid soils make up as much as 50% of the world’s soils, particularly in the tropics and subtropics where many developing countries are located. Also significant areas in the eastern and southern US have highly acidic soils. ARS researchers at the Robert W. Holley Center for Agriculture and Health at Ithaca, New York, discovered a unique protein in sorghum that allows sorghum to grow on acid soils. Knowing this, it will be possible to develop high yielding sorghum and other cereal crops to be grown on acid soils in developing countries including sub-Saharan Africa, Southeast Asia and South America, as well as on acid soils in the United States.
Yang, X., Yang, J., Pineros, M., Kochian, L.V., Zheng, S. 2011. Title A de novo synthesis citrate transporter VuMATE confers aluminum resistance in rice bean (vigna umbellata). Plant Cell and Environment. 34:2138-2148.
Ligaba, A., Maron, L., Shaff, J.E., Kochian, L.V., Pineros, M. 2012. Maize ZmALMT2 is a root anion transporter that mediates constitutive root malate efflux. Plant, Cell & Environment. 35(7):1365-3040.
Milner, M., Naoki, Y., Koyama, E., Jian, F., Kochian, L.V. 2012. Characterization of the high affinity Zn transporter from Noccaea caerulescens, NcZNT1, and dissection of its promoter for its role in Zn uptake and hyperaccumulation. New Phytologist. 195(1):113-123.
Caniato, F., Guimaraes, C., Hamblin, M., Billot, C., Rami, J., Maciel, B.H., Kochian, L.V., Liu, J., Garcia, A., Hash, C.T., Ramu, P., Mitchell, S., Kresovich, S., Oliveira, A., Avelar, G., Borem, A., Glaszmann, J., Schaffert, R.E., Magalhaes, J.V. 2011. The relationship between population structure and aluminum tolerance in cultivated sorghum. PloS One. 6(6):e20830. DOI: 10.1371/journal.pone.0020830.
Desousa, S., Clark, R.T., Mendes, F., De Oliveira, A., De Vasconcelos, M., Parentoni, S.N., Kochian, L.V., Guimaraes, C.T., Magalhaes, J.V. 2012. A role for root morphology and related candidate genes in P acquisition efficiency in maize. Functional Plant Biology. DOI: org/10.1071/FP12022.
Famoso, A., Zhao, K., Clark, R.T., Tung, C., Wright, M.H., Bustamante, C., Mccouch, S.R., Kochian, L.V. 2011. Genetic architecture of aluminum tolerance in rice (O. sativa) determined through genome-wide association analysis and QTL mapping. PLoS Genetics. 7(8):e1002221. DOI: 10.137/journal.pgen.1002221.
Liu, J., Luo, X., Shaff, J., Liang, C., Jia, X., Li, Z., Magalhaes, J., Kochian, L.V. 2012. A promoter swap strategy between the AtALMT and AtMATE genes increased arabidopsis aluminum resistance and improved carbon use efficiency for aluminum resistance. Plant Journal. 71(2):327-337.
Li, L., Yang, Y., Xu, Q., Owsiany, K., Welsch, R., Chitchumroonchokchai, C., Lu, S., Van Eck, J., Deng, X., Failla, M., Thannhauser, T.W. 2012. The Or gene enhances carotenoid accumulation and stability during post-harvest storage of potato tubers. Molecular Plant. 5:339-352.
Zhou, X., Fei, Z., Thannhauser, T.W., Li, L. 2012. Transcriptome analysis of ectopic chloroplast development in green curd cauliflower (Brassica oleracea L. var. botrytis). Biomed Central (BMC) Plant Biology. 11:169.