The metabolic expense of producing and operating neural tissue required for adaptive behaviour is considered a significant selective force in brain evolution. In primates, brain size correlates positively with group size, presumably owing to the greater cognitive demands of complex social relationships in large societies. Social complexity in eusocial insects is also associated with large groups, as well as collective intelligence and division of labour among sterile workers. However, superorganism phenotypes may lower cognitive demands on behaviourally specialized workers resulting in selection for decreased brain size and/or energetic costs of brain metabolism. To test this hypothesis, we compared brain investment patterns and cytochrome oxidase (COX) activity, a proxy for ATP usage, in two ant species contrasting in social organization. Socially complex Oecophylla smaragdina workers had larger brain size and relative investment in the mushroom bodies (MBs)-higher order sensory processing compartments-than the more socially basic Formica subsericea workers. Oecophylla smaragdina workers, however, had reduced COX activity in the MBs. Our results suggest that as in primates, ant group size is associated with large brain size. The elevated costs of investment in metabolically expensive brain tissue in the socially complex O. smaragdina, however, appear to be offset by decreased energetic costs.

My study investigated the evolution of oxidative metabolism in fishes. While intense selection for, or against, non-synonymous point mutations in coding sequence drives the evolution of mitochondrial OXPHOS genes, genome-specific mechanisms such as gene duplication events can play major roles in the evolution of nuclear OXPHOS genes. My thesis focused on the mitochondrial enzyme cytochrome c oxidase (COX), principally in fish because of their evolutionary origins and functional diversity in terms of energy metabolism. In the first part of my thesis, I examined a highly aerobic group of fishes (billfishes and tunas) to study the evolution of mitochondrial COX genes. Though the study began as a structure-function analysis of COX, my approach changed when my preliminary results called into question the accepted phylogenetic relationships of my species of interest. We generated a robust multigene phylogeny of this group to interpret data in a phylogenetically informative context. Phylogenetic analyses in this group provided us with a framework to study the evolution of mitochondrial OXPHOS genes, but unexpectedly revealed that: 1) billfishes are only distantly related to tunas, and share greater evolutionary affinities with flatfishes (Pleuronectiformes) and jacks (Carangidae), and 2) regional endothermy has evolved in a non-scombroid suborder in teleosts. These results collectively imply that regional endothermy has evolved independently at least twice within teleost fish. The second part of my thesis explored the evolution of the nuclear COX subunits, focusing on their origins in fish. Isoform transcription profiles coupled with phylogenetic analyses for each subunit show that vertebrate isoforms arose from a combination of early whole-genome duplications in basal vertebrates or specific lineages (e.g. teleosts), and more recent single gene duplication events. While there is evidence for retained function of some COX orthologues across fishes and mammals, others appear to have diverged in function since their earlier radiation, possibly contributing novel evolutionary functions. Together these two studies provide insight into the evolutionary forces facilitating adaptive change in mitochondrial and nuclear OXPHOS genes. / Thesis (Master, Biology) -- Queen's University, 2009-09-11 11:00:12.562

Spinobulbar muscular atrophy (SBMA) is a neurodegenerative disease caused by the expansion of a polyglutamine (polyGln) tract in the human androgen receptor (hAR). One mechanism by which polyGln-expanded proteins are believed to cause neuronotoxicity is through aberrant interaction(s) with, and possible sequestration of, critical cellular protein(s). / Our goal was to confirm and further characterize the interaction between hAR and cytochrome c oxidase subunit Vb (COXVb), a nuclear-encoded mitochondrial protein. We had previously isolated COXVb as an AR-interacting protein in a yeast two-hybrid search to identify candidates that interact with normal and polyGln-expanded AR. Using the mammalian two-hybrid system, we confirm that COXVb interacts with normal and mutant AR and demonstrate that the COXVb-normal AR interaction is stimulated by heat shock protein 70 (Hsp70). Also, BFP-tagged AR specifically co-localizes with cytoplasmic aggregates formed by GFP-labelled polyGln-expanded AR in androgen-treated cells. / Mitochondrial dysfunction may precede neuropathological findings in polyGln-expanded disorders and may thus represent an early event in neuronotoxicity. Interaction of COXVb and hAR, with subsequent sequestration of COXVb, may provide a mechanism for putative mitochondrial dysfunction in SBMA.

The biodiversity of Durban (eThekwini municipality) in KwaZulu Natal is primarily threatened by urbanization although other factors such as climate change and the spread of invasive species also pose a significant threat. Knowledge of what species exist within the city is important for biodiversity surveillance, detecting invasive taxa and uncovering cryptic species. Conducting a comprehensive biodiversity inventory is a daunting task, especially for hyperdiverse groups such as terrestrial arthropods, where closely related species can often only be separated by subtle morphological characters. This study investigated whether the barcoding marker, Cytochrome Oxidase C Subunit 1 (COI) can be used to efficiently and accurately delineate species of ants (family Formicidae) in comparison to traditional taxonomic approaches. The feasibility of DNA barcoding for assembling biodiversity inventories for urban areas which could be useful in conservation planning was also evaluated. A total of 619 individuals were sequenced from 23 geographic localities within the eThekwini region and surrounding regions. DNA barcoding revealed 80 provisional species/ “barcode clusters” or monophyletic lineages which could represent distinct species, while morphology revealed 51 different morphospecies. Extrapolation measures of species richness indicated that as many as 153 species of ants could occur in the city. Phylogenetic and phylogeographic analyses were performed on co-distributed species belonging to the genera Lepisiota, Camponotus, Pheidole and Pachycondyla to better understand the spatial distribution of genetic variability in the eThekwini region. Nuclear markers 18S rDNA and 28S rDNA were also sequenced and compared for a subsample of individuals from Camponotus and Pachycondyla. There was genetic variation at COI and the nuclear markers for each of the species examined. In order to fully elucidate the population genetic patterns which could be expected in eThekwini and surrounding regions, further sampling across more localities is essential. The use of more nuclear markers could also assist in uncovering these unique patterns of genetic variation in an urban setting. In this study, the utility of COI as a species diagnostic tool in ants was confirmed. The barcoding library constructed showed promise in highlighting reserves that should be preserved and possible cryptic speciation for further investigation. / M. Sc. University of KwaZulu-Natal, Durban 2014.

This thesis describes a study of the substrate selectivity of active site mutants of the monooxygenase cytochrome P450_cam. A range of mutants was constructed which replaced the phenolic side-chain at the Tyr-96 position by various hydrophobic amino acid residues. These 'hydrophobic mutants' were then combined with other mutations around the active site (Val-247, Phe-87, Ile-395 and Phe-193) which altered the space available at different positions in the active site. These mutants were then tested with an in vitro reconstituted P450_cam system with a range of substrates related to diphenylmethane and phenylcylcohexane. All of these large compounds were poor substrates for the wild-type enzyme. It was found that it was necessary to increase both the space available in the active site and the active site hydrophobicity to achieve substrate turnover. The substrates were oxidised preferentially on the aliphatic cyclohexyl ring over the more constrained phenyl ring suggesting that the active site is predisposed to binding the cyclohexyl ring close to the haem. Hydroxylation using the in vitro reconstituted P450_cam system is limited by catalyst lifetime and the need for the expensive cofactor NADH. For P450_cam hydroxylation to become a viable synthetic method it is necessary to find ways to bypass the use of NADH. For this reason various self-sufficient P450_cam system were constructed and expressed in E. coli. The best of these, despite limited protein expression, was found to turnover camphor with the wild-type P450_cam enzyme and other substrates with the Y96A mutant. The in vivo catalytic system was then used to screen many P450_cam mutants for the oxidation of natural products, monoterpenes and sesquiterpenes (e.g. limonene, pinene and valencene). Most of the target substrates are not oxidised by the wild-type enzyme but all are hydroxylated by some if not all of the P450_cam mutants with different degrees of selectivity. Some of the products identified so far are important compounds in the field of flavour and fragrance chemistry (e.g. verbenol and nookatone).