Amino acids (AA) are the building blocks of proteins, which makes them extremely important for growth and development. Crop seeds, such as legumes and cereals, play an essential role as a key food source in the diet of humans and livestock but do not meet the dietary requirements of essential amino acids (EAA), which are the AA that humans and vertebrates cannot synthesize and must obtain from the diet. Lacking sufficient levels of EAA in the diet can lead to protein-energy malnutrition, which adversely affect the immune, gastrointestinal, nervous, and cardiovascular systems. Efforts to fortify AA composition in crop seeds have had a very limited success since plants respond to induced protein composition alterations by activating a regulatory mechanism that “resets” it back to the original state. This phenomenon is known as proteome re-balancing, and, while beneficial for plants’ growth and development, has been a major hurdle to biofortification. A good understanding of the amino acid composition regulation and rebalancing mechanism in seeds would improve the biofortification of AA composition.

Chapter One provides a comprehensive introduction from previous and what is known about amino acid composition in the seeds, the challenges identified in previous experimentation, and how the content of the other chapters builds upon and adds value to the area of seed amino acid research and to the understanding of proteome rebalancing. Chapter Two emphases on uncovering the genes and biological processes that underly proteome relancing using the mutants from the first most abundant seed storage proteins (SSPs), the cruciferins also known as 12S. To better understand how proteome rebalancing is achieved in seeds, we conducted a comprehensive analysis on an Arabidopsis mutants lacking the three most abundant SSPs, the cruciferins (CRUs). The multi-omics analysis such as transcriptome, proteome, metabolome, and measurement of physiology parameters  was conducted on singles (crua, crub, and cruc) and the triple know out (cruabc) mutants and compared them to the wild type (Col-0). All major seeds storage compounds remained unchanged in these mutants suggesting rebalanced seeds. further analysis show that translation and oxidative stress responses are the two key biological processes that dominated the proteome rebalancing. Chapter Three focuses on how proteome rebalancing is achieved in the second most abundant seed storage proteins known as the Napins known as 2S. In this chapter, I used only one mutant, which is napin-RNAi (RNA interference construct that target all 5 member genes of this group). Using multi-omics analyses, we found that all reserve compounds remained the same compared to Col-0 expect the sulfur. Only oxidative stress response dominated the biological processes involved in proteome rebalancing in the Napin seed storage proteins. Chapter Four covered the remained seed storage protein mutants from the cruciferins, these were the doubles (cruab, cruac, and crubc). In this chapter, all analyses done compared the doubles (cruab, cruac, and crubc) to Col-0; genes and key pathways involved in proteome rebalancing were revealed. Lastly, Chapter Five the conclusion, reiterates the contributions of this dissertation to the field of seed amino acid proteome rebalancing and provides future directions and research projects that can be done to bring much more insight in this field.

Publications

Bagaza C., Ansaf H., Yobi A., Chan Y.O., Slaten M.L., Czymmek K., Joshi T., Mittler R., Mawhinney T.P., Cohen D.H., Yasuor H., Angelovici R. A multi-omics approach reveals a link between ribosomal protein alterations and proteome rebalancing in Arabidopsis thaliana seeds (2024) Plant Journal, 120(6), 2803-27.

Katz E., Li J.-J., Jaegle B., Ashkenazy H., Abrahams S.R., Bagaza C., Holden S., Pires C.J., Angelovici R., Kliebenstein D.J. Genetic variation, environment and demography intersect to shape Arabidopsis defense metabolite variation across Europe (2021) eLife, 10, art. no. e67784

Slaten M.L., Yobi A., Bagaza C., Chan Y.O., Shrestha V., Holden S., Katz E., Kanstrup C., Lipka A.E., Kliebenstein D.J., Nour-Eldin H.H., Angelovici R. MGWAS uncovers Gln-glucosinolate seed-specific interaction and its role in metabolic homeostasis (2020) Plant Physiology, 183(2), 483-500. 

Yobi A., Bagaza C., Batushansky A., Shrestha V., Emery M.L., Holden S., Turner-Hissong S., Miller N.D., Mawhinney T.P., Angelovici R. The complex response of free and bound amino acids to water stress during the seed setting stage in Arabidopsis (2020) Plant Journal, 102(4), 838-55. 

Bagaza C., Ansaf H., Yobi A., Chan Y.O., Slaten M.L., Czymmek K., Joshi T., Mittler R., Mawhinney T.P., Cohen D.H., Yasuor H., Angelovici R. A multi-omics approach reveals a link between ribosomal protein alterations and proteome rebalancing in Arabidopsis thaliana seeds (2024) Plant Journal, 120(6), 2803-27

Li M., An H., Angelovici R., Bagaza C., Batushansky A., Clark L., Coneva V., Donoghue M.J., Edwards E., Fajardo D., Fang H., Frank M.H., Gallaher T., Gebken S., Hill T., Jansky S., Kaur B., Klahs P.C., Klein L.L., Kuraparthy V., Londo J., Migicovsky Z., Miller A., Mohn R., Myles S., Otoni W.C., Pires J.C., Rieffer E., Schmerler S., Spriggs E., Topp C.N., Van Deynze A., Zhang K., Zhu L., Zink B.M., Chitwood D.H. Topological data analysis as a morphometric method: Using persistent homology to demarcate a leaf morphospace (2018) Frontiers in Plant Science, 9, art. no. 553. 

Doctoral Program Committee

• Dr. Ruthie Angelovici, Chair
• Dr. Paula McSteen 
• Dr. Sherry Flint-Garcia
• Dr. Bing Yang