Progress 09/01/07 to 08/31/08
Outputs
OUTPUTS: -- We are testing the hypothesis that sorbitol dehydrogenase (SDH) and aldose reductase (AR) could contribute to a centrally-important role for sugar alcohols in development of maize kernels and possibly other grains. Kernels do not import or store sorbitol, so an intermediary, cycling, or short-distance transport function appears likely. To address this possibility, we have identified-, and are now characterizing-, a maize mutant deficient in the sole gene for sorbitol dehydrogenase (Sdh1). This mutation reduces the maximal SDH activity in developing kernels to less than 6% of wild-type levels. The resulting phenotype is a 20% smaller kernel under field conditions (dried-seed weight at maturity, significant to p<.001). The sdh1 mutation also increased sugar levels during development, especially at 25 days after pollination, near the harvest date for most sweet corn lines. At this stage, hexose levels more than doubled, and sucrose levels were elevated by 16%. A central role for sorbitol in the sugar balance of developing kernels was thus indicated. The sorbitol levels themselves were reduced by only 33% in the mutant kernels at this stage of development, indicating that SDH may not be the only sorbitol-handling enzyme in the maize kernel. Aldose reductase (AR) is also a candidate, but structural and kinetic analysis of AR genes expressed in kernels indicate that they are likely to have a less active role (de Sousa et al. 2008). -- We have also able to enhance cost and specificity of expression profiles by developing a 3'UTR-expression profiling method (Eveland et al. 2008). Information returned per unit cost was increased by 1) targeting all reads to the gene-specific, 3'UTR, and 2) concurrently sequencing multiple samples in a single, pooled run (sequences were assigned to their original sources using DNA barcodes incorporated into sub-libraries at the time of construction). Specificity rose markedly, since the 3'UTR distinguished not only gene-specificity among gene families, but also allele specificity. Instrumental to this approach was the relatively long-read technology associated with the 454-based sequencing approach. This method also allowed quantification of expression across three-orders of magnitude, and detection of transcripts not present on arrays. -- Thus far we have used this method to successfully delineate gene family members most responsive to genetic or developmental changes. We have focused our efforts on sink structures, primarily developing ovaries of mutant and wild-type plants, and also on root hairs. In each of these efforts, specific genes have emerged for cell-wall biosynthesis, C/N balance, and phenylpropanoid biosynthesis. PARTICIPANTS: Not relevant to this project. TARGET AUDIENCES: Not relevant to this project. PROJECT MODIFICATIONS: Not relevant to this project.
Impacts
-- The sdh1 mutant continues to offer insights into potential roles of sorbitol in grain development. -- The sdh1 mutant also has potential as a new sweetcorn, and may be useful in sweetcorn breeding efforts. -- The 3'UTR 454 profiling method provides a less-expensive and more specific means of expression profiling. The capacity to individually quantify expression of gene-family members can be invaluable to efforts to dissect their functional roles (in photosynthate partitioning or other processes).
Publications
- Koch, K.E., de Sousa, S.M. (2008) Material and methods for modulating seed size, seed number, and seed sugar content in plants. International Application No. PCT/US2008/064130; filed May 19, 2008 UF#-12553.
- de Sousa, S.M., Rosselli, L. K., Kiyota, E., da Silva, J.C., Souza, G.H.M.F., Peroni, L.A., Stach-Machado, D.R., Eberlin, M., Souza, A.P., Koch, K.E., Arruda, P., Torriani, I.L., Yunes, J.A. (2008). Structural and kinetic characterization of a maize aldose reductase, Plant Physiology and Biochemistry. 47: 98-104.
- Eveland,A.L., McCarty, D.R., Koch, K.E. (2008). Transcript profiling by 3'-Untranslated region sequencing resolves expression of gene families. Plant Physiology. 146: 32-44.
- de Sousa, S.M., Hunter III, C.T., Ankumah, N., Avigne, W.T., Koch, K.E. (2008) Sorbitol Accumulation is elevated during the development of maize kernels deficient in sorbitol dehydrogenase, Annual Meeting of the American Society of Plant Biologists, Merida, Yucatan, Mexico.
- de Sousa, S.M., Hunter III, C.T., Ankumah, N., Avigne, W.T., Koch, K.E. (2008) A sorbitol dehydrogenase deficiency increases sugar levels during maize development, 50th Annual Maize Genetics Conference, Washington, DC, USA.
- de Sousa, S.M. (2008) Sugar accumulation is elevated during the development of maize kernels deficient in sorbitol dehydrogenase, 19th Annual Plant Molecular and Cellular Biology Program (PMCB), Jacksonville Beach, FL, USA. Workshop speaker.
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