Characterization of land plant-specific proteins required for mitochondrial translation initiation

Plant mitochondria produce the majority of adenosine triphosphate (ATP) – the cellular energy currency for metabolic reactions needed for plant growth, development, and maintenance. Despite a relatively thorough understanding of basic mitochondrial functions, many mitochondrial proteins and processes remain poorly understood. The aims of this thesis are to i) review and compare the mitochondrial unfolded protein response (UPRmt) and related signalling across eukaryotic kingdoms, to ii) describe an efficient isolation method of Arabidopsis mitochondria using continuous Percoll density gradients, and to iii) characterize two Arabidopsis thaliana genes, mTRAN1 and mTRAN2. Paper I summarizes the current knowledge of UPRmt across eukaryotic kingdoms, and describes a meta-analysis of UPRmt regulators and target genes. UPRmt is a mitochondria-to-nucleus “retrograde” response that regulates nuclear gene expression during mitochondrial dysfunction to maintain mitochondrial homeostasis. Although UPRmt has been extensively studied in animals, relatively little is known about the plant UPRmt and only few regulators have recently been identified. In yeast, very few unfolded protein responses that seem to be related to UPRmt have been described. Here, the UPRmt in animals, yeast and plants are compared. Our study indicates that each kingdom has evolved their own specific regulators, which however induce very similar groups of target genes. Our meta-analysis identifies homologs of known UPRmt regulators and responsive genes across eukaryotic kingdoms.Paper II describes a strategy for efficient purification of Arabidopsis mitochondria using continuous Percoll density gradients. By using this method, the purity of isolated mitochondria is greatly improved. Obtained mitochondria can be either used for assays requiring highly intact and functional mitochondria, e.g. import assay or respiration measurement, or be stored for later use, e.g. BN-PAGE or western blot.Paper III describes th