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Recently, we demonstrated that the anti-bacterial agent tigecycline preferentially induces death in leukemia cells through the inhibition of mitochondrial protein synthesis. To better understand the mechanisms of sensitivity and resistance to mitochondrial translation inhibition, we treated TEX leukemia cells with increasing concentrations of tigecycline over 4 months, and selected a population of cells resistant to tigecycline (RTEX+TIG). Compared to their wild type counterparts, the resistant RTEX+TIG cells had an altered metabolic profile with diminished oxidative phosphorylation and a greater reliance on glycolysis. Upon removal of tigecycline from RTEX+TIG cells, the cells re-established aerobic metabolism and oxidative phosphorylation to wild type levels. At the molecular level, these cells had increased levels of HIF1a. Strikingly, tigecycline-resistant cells had decreased expression of CD34 and CD117, clonogenic growth potential and engraftment capabilities in vivo. Thus, chronic inhibition of mitochondrial translation leads to the establishment of rho-zero-like metabolic phenotype, and is associated with differentiation of leukemia cells.

The focus of this thesis has been the phylogenomics and evolution of the Alphaproteobacteria. This is a very diverse group which encompasses bacteria from intraceullar parasites, such as the Rickettsiales, to freeliving bacteria such as the most abundant bacteria on earth, the SAR11. The genome sizes of the Alphaproteobacteria range between 1 Mb and 10 Mb. This group is also connected to the origin of the mitochondria. Several studies have placed the SAR11 clade together with the Rickettsiales and mitochon- dria. Here I have shown that this placement is an artifact of compositional heterogeneity. When choosing genes or sites less affected by heterogeneity we find that the SAR11-clade instead groups with free-living alphaproteobacteria. Gene-content analysis showed that SAR11 was missing several genes for recombination and DNA-repair. The relationships within the SAR11- clade has also been examined and questioned. Specifically, we found no support for placing the taxon referred to as HIMB59 within the SAR11. Ocean metagenomes have been investigated to determine whether the SAR11-clade is a potential relative of the mitochondria. No such relationship was found. Further I have shown how important it is to take the phylogenetic relationships into account when doing statistical analyzes of genomes. The evolution of LD12, the freshwater representative of SAR11, was investigated. Phyloge- nies and synonymous substitution frequencies showed the presence of three distinct subclades within LD12. The recombination to mutation rate was found to be extremely low. This is re- markable in light of the very high rate in the oceanic SAR11. This is may be due to adaptation to a more specialized niche. Finally we have compared structure-based and sequence-based methods for orthology pre- diction. A high fraction of the orfan proteins were predicted to code for intrinsically disordered proteins. Many phylogenetic methods are sensitive to heterogeneity and this needs to be taken into ac- count when doing phylogenies. There have been at least three independent genome reductions in the Alphaproteobacteria. The frequency of recombination differ greatly between freshwater and oceanic SAR11. Forces affecting the size of bacterial genomes and mechanisms of evolu- tionary change depend on the environmental context.