Studies of E2s Related to Unconventional Ubiquitination in Arabidopsis thaliana
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Protein ubiquitination, a fundamental protein modification cascade, is catalyzed by three types of enzymes referred to as E1 for ubiquitin (Ub) activation, E2 for Ub conjugation and E3 for Ub ligation. Arabidopsis has 37 E2 genes and functions for most of them remain largely unknown. Further, although the conventional Lys48-linked protein ubiquitination is well known for its function in protein degradation, functions for protein ubiquitination linked through other lysine residues of Ub, e.g. Lys11 and Lys63 are poorly understood in plants. This study aims to advance the understanding of the functions of Arabidopsis E2s involved in the unconventional ubiquitination. Previously, UBC13A/B have been identified as the E2 for Lys63-linked polyubiquitination, and ubc13 mutant plants display strong phenotypes in root development. In this study, a family of six E3s that can interact with UBC13 were identified, and when a subgroup of three genes were inactivated, the T-DNA mutants showed a glucose-sensitive phenotype suggesting their possible role in sugar signaling. It has been proposed that in root development UBC13 may control the level of Aux/IAA (Auxin/indole-3-acetic acid) proteins which regulate the activity of auxin response factors (ARFs). Several ARFs were evaluated, and overexpression of ARF6 enhanced the root hair growth in WT plants and partially rescued the root phenotypes of the ubc13 mutant. A major part of this study was on Arabidopsis UBC22, a sole member of an Arabidopsis E2 subfamily. Results from the phylogenetic analysis and in vitro ubiquitination assay indicate that UBC22 is an E2 for Lys11-linked ubiquitination in plants. The ubc22 knockout mutant had much reduced seed setting, and further analysis showed that UBC22 plays a critical role in the formation of functional megaspore and female gametophyte. Moreover, certain ubc22 mutant plants showed distinct phenotypes in vegetative development. The RNA-seq analysis revealed that genes of different pathways were differentially affected in different subtypes of plants. Plants of one subtype had increased expression of genes in pathogen defense and showed an enhanced resistance to a plant pathogen. Therefore, the present results have provided a new understanding on the function of UBC13, identified UBC22 as a novel E2, and revealed its multiple functions in plants.
DegreeDoctor of Philosophy (Ph.D.)
CommitteeDatla, Raju; Lee, Jeremy; Wei, Yangdou; Wu, Yuliang
Copyright DateDecember 2018