posted on 2022-03-28, 19:08authored byAnte Jerkovic
Abscisic acid (ABA) is a well known plant hormone that is involved in many biotic and abiotic stress responses. Application of ABA in the milling of wheat has been shown to improve flour yield and quality. This suggests that there may be biochemical processes which impart a physiological change that is favorable to improving milling performance. In this study, I have attempted to better understand the biochemical and physiological changes of water + ABA-conditioned grain. The strategy involved four stages: (1) apply proteomic analysis to measure and compare the differential expression of proteins in the germ, bran (aleurone layer) and ventral groove of non-conditioned grain (control) and grain conditioned with water-and water + ABA, (2) localize proteins of interest that were identified in the proteomic analysis using immunolabeling and confocal microscopy, (3) test for flour yield and quality by analysing variations in grain fractionation using a laboratory scale mill (MicroMill) to generate fractionation quality scores (FQS's) (4) measure physiological changes in water relations by determining grain water potential (ψ) and mechanical properties of the germ and bran/endosperm of following treatments. The proteomic analysis of the water-conditioned grain showed that there were many changes in protein levels in the bran and ventral groove tissues, however, there were almost no changes in protein levels in the germ tissue. In the tissues of the ABA-conditioned grain, there were many differentially expressed proteins in the germ, bran and ventral groove tissues, especially those involved in biotic and abiotic stress response. Of these proteins, two are involved in water relations; late embryogenesis abundant proteins (LEA's) and tonoplast intrinsic protein's (TIP's). The ventral groove showed little variation in protein levels between water-and water + ABA-conditioned grain; however, both exhibited a ~ 5-fold increase in LEA's compared to non-conditioned grain. LEA protein abundance also increased in the germ and aleurone cells after ABA treatment but only by ~ 1.5-fold. Confocal microscopy of immunolabeled grain cross-sections, revealed that group 2 LEA (dehydrin) proteins are distributed throughout the intracellular matrix in a 'honeycomb-like' arrangement and also surrounding the nucleus and inner cell walls within the germ cells, aleurone cells in the bran layer and in the aleurone cells surrounding the ventral groove. TIP levels decreased by ~ 2-fold exclusively in the germ, which is likely to reduce water movement in and out of the tonoplast. An increase in FQS's of ABA-conditioned grain compared with water-conditioned grain may indicate improved fractionation, leading to improved flour quality and yield. Psychrometric measurements of the germ-end and bran/endosperm-end of ABA-conditioned grain showed a slightly elevated ψ when compared to water-only treatment after drying at room temperature for 1.5 h. This suggested that the grain somehow retains more moisture if treated with ABA. Finally, mechanical property analysis of the germ and bran tissue showed that the germ was softer and bran was tougher after conditioning with ABA. Collectively, these results suggest that ABA may induce changes in biochemical processes of the germ and aleurone cells of the bran and ventral groove tissues, such as increased levels of LEAs and a reduction in TIPs in order to prevent moisture loss in response to an environmental stress such as drought. Consequently it is suggested that this altered the water distribution within the grain, thus transforming its physiological properties, making it better adapted to surviving a desiccating environment. From an applied science perspective, this physiological change allows the grain to be more amenable to milling, thus improving flour yield and quality.
Table of Contents
1. Introduction -- 2. Strategic distribution of protective proteins within bran layers of wheat (Triticum aestivum L.) protects the nutrient-rich endosperm -- 3. Proteomic analysis of water-and water + abscisic acid (ABA)-conditioned grain -- 4. iTRAQ analysis of bran and ventral groove tissues from non-conditioned grain -- 5. Identification and immunolocalisation microscopy of late embryogenesis abundant (LEA) proteins -- 6. Grain fractionation and physiological analysis of water-and water + ABA-conditioned wheat grain -- 7. Conclusion and future direction.
Includes bibliographical references
Thesis submitted in fulfilment of the requirements for the degree of Doctor of Philosophy, Macquarie University, Dept. of Chemistry and Biomolecular Sciences, 2011.
Thesis (PhD), Macquarie University, Faculty of Science, Dept. of Chemistry and Biomolecular Sciences
Department, Centre or School
Department of Chemistry and Biomolecular Sciences
Year of Award
Copyright Ante Jerkovic 2011.
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