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Open Access Thesis
Master of Science (M.S.)
Year Degree Awarded
Month Degree Awarded
Mitochondria play critical roles not only in primary metabolism as a central organelle for ATP generation, but also in responding to abiotic stresses. We identified a mutation in the MTERF18 (Mitochondrial Transcription Termination factor)/SHOT1 (Suppressor of hot1-4 1) gene in Arabidopsis thaliana that enables plants to better tolerate heat and oxidative stresses, presumably due to reduced oxidative damage, but the exact molecular mechanism of the heat tolerance is unknown. In order to reveal the stress tolerance mechanisms of mterf18/shot1 mutations, it is critical to understand the molecular defects of the mutant and to identify the molecular targets of the MTERF18/SHOT1 protein. MTERF18/SHOT1, a mitochondrial matrix protein, was found to bind to membrane-spanning mitochondrial AAA+ proteins homologous to ATAD3a of humans and other multicellular eukaryotes. A. thaliana has four ATAD3a homologues in two clades, and plants require one gene from each clade for viability. Previous studies of the topology and ATPase activity of ATAD3a suggest a role in endoplasmic reticulum (ER)-mitochondria contact sites. These sites are poorly defined in plants, and their relationship to heat stress tolerance is intriguing. To better understand ATAD3 function I expressed and purified the soluble, matrix-located, catalytic C-terminal ATPase domain of these proteins in order to assay their ATPase activity and oligomerization states. Transgenic plants with fluorescently labelled ER and mitochondria have been generated to observe effects of the MTERF18/SHOT1 mutation on ER-mitochondria dynamics. These studies of the four ATAD3 proteins will provide insights into ER-mitochondrial contact sites in plants, and into their link to MTERF18/SHOT1 and heat stress tolerance. I also provide a review of our current knowledge of ER-mitochondria contact site protein components in plants with reference to these proteins in A. thaliana.
Table 2.1- pCR8 Vectors.xlsx (71 kB)
Table 2.2- pET23b His-SUMO vectors .xlsx (53 kB)
Table 2.3- BiFC vectors.xlsx (72 kB)
Table 4.1. ER-rb-transformed plant lines.xlsx (10 kB)
Table 5.1- Yeast-specific ERMES and MICOS subunits.xlsx (10 kB)
Table 5.2. The known plant ERMIONE complex and its constituent subcomplexes and proteins.xlsx (16 kB)
Zelman, Sam, "Investigation of SHOT1-binding ATPases in Arabidopsis thaliana" (2020). Masters Theses. 1000.