Evolutionary Genomic Responses to Mitochondrial Dysfunction in Caenorhabditis elegans

Understanding the impact of mitochondrial dysfunction on genome evolution has the potential not only to provide new insights on the basic evolutionary processes influencing mitochondrial and nuclear genomes, but may also reveal novel avenues for evolutionary adaptive recovery from harmful mutations. Aberrant mitochondrial activity is fundamental to the pathology of mitochondrial diseases in addition to neurodegenerative disorders. While the effects of mitochondrial dysfunction have received much attention, less is known about their impact on genome evolution and potential target mechanisms for ameliorating the harmful effects of mitochondrial impairment. Characterizing genome modifications in animal populations predisposed to mitochondrial dysfunction may identify novel genes, mechanisms, and physiological pathways to target for recovery and provides a genome-wide perspective on the impact of aberrant mitochondrial activity. This dissertation research investigates how mitochondrial and nuclear genomes evolve in organisms genetically predisposed to mitochondrial dysfunction and contrasts genomic evolution in large and small population sizes. This work furthers understanding of the impact of evolutionary forces which influence genome evolution in population with reduced fitness, and reveals new insights into genomic responses to mitochondrial dysfunction. Chapters 2 and 3 of this dissertation focus on genome evolution using a set of mitochondrial respiratory chain mutant (gas-1¬ strain) and wild-type (N2 strain) Caenorhabditis elegans mutation-accumulation (MA) lines that experienced single-worm bottlenecking. The N2 MA lines, derived from a previous experiment, were bottlenecked for 250 generations. The gas-1 MA lines were created for this research, and bottlenecked in the laboratory for a maximum of 50 generations. Chapter 2 investigates mitogenomic evolution and heteroplasmic inheritance patterns evolving under extreme drift in gas-1 and N2 MA lines. Chapter 3 analyzes nuclear genome evolution using this same set of gas-1 and N2 MA lines. In contrast, Chapter 4 provides a complementary perspective, analyzing mitochondrial and nuclear genome evolution in twenty-four gas-1 'recovery line' (RC) populations, evolved in large population sizes for sixty generations. Bioinformatic methods and computational simulations were applied to characterize and evaluate genome evolution and provide a comprehensive investigation of the impact of mitochondrial dysfunction within a population genetics framework. In Chapter 2 our results of inherited mitochondrial DNA (mtDNA) heteroplasmy are in alignment with predictions of theories where a small subset of mtDNA molecules from the parental generation repopulates the mitochondrial genome pool for the progeny. Comparisons between Chapter 2 and 4 suggest that in both gas-1 and N2 strains organelle genome copy number is elevated in an environment characterized with extreme genetic drift but is less impacted throughout evolution in large populations when the force of genetic drift is reduced. Investigation of nuclear genome evolution in Chapter 3 revealed putative beneficial nuclear mutations in bottlenecked gas-1 populations. Additionally, compared to the N2 MA lines, the gas-1 MA lines were also observed to have a greater number of mutations located within the gas-1 gene interaction network. These observations reveal new insights into the potential fitness landscape for beneficial mutation and how nuclear genome evolution differs when predisposed to mitochondrial dysfunction in an environment characterized by extreme genetic drift. In Chapter 4, focusing on evolution in large populations, we observed parallel and potentially compensatory mitochondrial mutations indicative of positive selection in the gas-1 RC lines. Identified at heteroplasmy levels near-fixation, these mtDNA mutations were located in genes predicted to physically interact with the gas-1 gene. As signatures of positive selection were not detected in the mitochondrial genomes of gas-1 MA lines analyzed in Chapter 2, this work suggests that the processes by which beneficial mtDNA mutations rise to homoplasmy within the population may be less likely to occur in small populations. Additionally, we determined the evolutionary rate of nuclear genome change in Chapter 4 to be three times slower than published mutations rate values for C. elegans suggesting the influence of purifying selection in RC lines. Given that a quarter of nuclear mutations were located in genes exhibiting interactions within two-degrees of gas-1 it is likely that positive selection also influenced nuclear genome evolution. Overall, this research demonstrates that although adaptation from harmful mutation may occur in small or large populations, the observed paths to evolutionary adaptive recovery involve different mechanisms and suggests that although an environment with pervasive genetic drift may permit the fixation of beneficial nuclear mutations, the processes by which beneficial mtDNA mutations rise to homoplasmy within the population may be less permissive

Selfish Mitochondrial DNA Proliferates and Diversifies in Small, but not Large, Experimental Populations of Caenorhabditis briggsae

Evolutionary interactions across levels of biological organization contribute to a variety of fundamental processes including genome evolution, reproductive mode transitions, species diversification, and extinction. Evolutionary theory predicts that so-called “selfish” genetic elements will proliferate when the host effective population size (Ne) is small, but direct tests of this prediction remain few. We analyzed the evolutionary dynamics of deletion-containing mitochondrial DNA (ΔmtDNA) molecules, previously characterized as selfish elements, in six different natural strains of the nematode Caenorhabditis briggsae allowed to undergo experimental evolution in a range of population sizes (N = 1, 10, 100, and 1,000) for a maximum of 50 generations. Mitochondrial DNA (mtDNA) was analyzed for replicate lineages at each five-generation time point. Ten different ΔmtDNA molecule types were observed and characterized across generations in the experimental populations. Consistent with predictions from evolutionary theory, lab lines evolved in small-population sizes (e.g., nematode N = 1) were more susceptible to accumulation of high levels of preexisting mtDNA compared with those evolved in larger populations. New ΔmtDNA elements were observed to increase in frequency and persist across time points, but almost exclusively at small population sizes. In some cases, mtDNA levels decreased across generations when population size was large (nematode N = 1,000). Different natural strains of C. briggsae varied in their susceptibilities to mtDNA accumulation, owing in part to preexisting compensatory mtDNA alleles in some strains that prevent deletion formation. This analysis directly demonstrates that the evolutionary trajectories of ΔmtDNA elements depend upon the population-genetic environments and molecular-genetic features of their hosts

Microwave-induced control of Free Electron Laser radiation

The dynamical response of a relativistic bunch of electrons injected in a planar magnetic undulator and interacting with a counterpropagating electromagnetic wave is studied. We demonstrate a resonance condition for which the free electron laser (FEL) dynamics is strongly influenced by the presence of the external field. It opens up the possibility of control of short wavelength FEL emission characteristics by changing the parameters of the microwave field without requiring change in the undulator's geometry or configuration. Numerical examples, assuming realistic parameter values analogous to those of the TTF-FEL, currently under development at DESY, are given for possible control of the amplitude or the polarization of the emitted radiation.Comment: 14 pages, 5 figures, accepted for publication in Phys. Rev.

Electrical behaviour, characteristics and properties of anodic aluminium oxide films coloured by nickel electrodeposition

Porous anodic films on 1050 aluminium substrate were coloured by AC electrodeposition of nickel. Several experiments were performed at different deposition voltages and nickel concentrations in the electrolyte in order to correlate the applied electrical power to the electrical behaviour, as well as the characteristics and properties of the coatings. The content of nickel inside the coatings reached 1.67 g/m2, depending on the experimental conditions. According to the applied AC voltage in comparison with the threshold voltage Ut, the coating either acted only as a capacitor when U\Ut and, when U[Ut, the behaviour during the anodic and cathodic parts of the power sine wave was different. In particular, due to the semi-conducting characteristics of the barrier layer, additional oxidation of the aluminium substrate occurred during the anodic part of the electrical signal, whilst metal deposition (and solvent reduction) occurred during the cathodic part; these mechanisms correspond to the blocked and pass directions of the barrier layer/electrolyte junction, respectively

The celebrity entrepreneur on television: profile, politics and power

This article examines the rise of the ‘celebrity entrepreneur’ on television through the emergence of the ‘business entertainment format’ and considers the ways in which regular television exposure can be converted into political influence. Within television studies there has been a preoccupation in recent years with how lifestyle and reality formats work to transform ‘ordinary’ people into celebrities. As a result, the contribution of vocationally skilled business professionals to factual entertainment programming has gone almost unnoticed. This article draws on interviews with key media industry professionals and begins by looking at the construction of entrepreneurs as different types of television personalities and how discourses of work, skill and knowledge function in business shows. It then outlines how entrepreneurs can utilize their newly acquired televisual skills to cultivate a wider media profile and secure various forms of political access and influence. Integral to this is the centrality of public relations and media management agencies in shaping media discourses and developing the individual as a ‘brand identity’ that can be used to endorse a range of products or ideas. This has led to policy makers and politicians attempting to mobilize the media profile of celebrity entrepreneurs to reach out and connect with the public on business and enterprise-related issues

Quantitative imaging of coronary blood flow

Positron emission tomography (PET) is a nuclear medicine imaging modality based on the administration of a positron-emitting radiotracer, the imaging of the distribution and kinetics of the tracer, and the interpretation of the physiological events and their meaning with respect to health and disease. PET imaging was introduced in the 1970s and numerous advances in radiotracers and detection systems have enabled this modality to address a wide variety of clinical tasks, such as the detection of cancer, staging of Alzheimer's disease, and assessment of coronary artery disease (CAD). This review provides a description of the logic and the logistics of the processes required for PET imaging and a discussion of its use in guiding the treatment of CAD. Finally, we outline prospects and limitations of nanoparticles as agents for PET imaging

Spinocerebellar ataxia type 6 family with phenotypic overlap with Multiple System Atrophy

Aim of the study. Multiple system atrophy (MSA) and spinocerebellar ataxia (SCA) share similar symptomatology. We describe a rare occurrence of familial MSA that proved to be SCA6 upon genetic analysis.Materials and methods. Eighty MSA patients were enrolled in our study; blood samples were collected and genetic screening of the familial case for known SCA loci was performed.Results. A 68-year-old woman presented with recurrent and severe episodes of light-headedness, imbalance, frequent falls, neck and lower back stiffness, subjective arm and leg weakness, and numbness and tingling in both feet. One year later, her condition had declined; she experienced more falls, worsening instability, again more generalised but still subjective weakness, impaired fine motor movements, slurred speech, difficulty swallowing, episodes of choking, bladder incontinence, and constipation. Clinical suspicion included parkinsonism, MSA, and SCA. The patient was enrolled in our MSA study and was found to have 22 and 12 CAG repeats in CACNA1A. The other 79 clinical MSA patients were negative for SCA6 screening.Conclusions and clinical implications. While MSA and SCA may have similar presentations during early disease stages, the presence of both conditions on the list of differential diagnoses can be a diagnostic dilemma. Further analysis will aid in developing a biomarker to distinguish between the two conditions and guide proper management

Protein aggregation and calcium dysregulation are hallmarks of familial Parkinson's disease in midbrain dopaminergic neurons

Mutations in the SNCA gene cause autosomal dominant Parkinson’s disease (PD), with loss of dopaminergic neurons in the substantia nigra, and aggregation of α-synuclein. The sequence of molecular events that proceed from an SNCA mutation during development, to end-stage pathology is unknown. Utilising human-induced pluripotent stem cells (hiPSCs), we resolved the temporal sequence of SNCA-induced pathophysiological events in order to discover early, and likely causative, events. Our small molecule-based protocol generates highly enriched midbrain dopaminergic (mDA) neurons: molecular identity was confirmed using single-cell RNA sequencing and proteomics, and functional identity was established through dopamine synthesis, and measures of electrophysiological activity. At the earliest stage of differentiation, prior to maturation to mDA neurons, we demonstrate the formation of small β-sheet-rich oligomeric aggregates, in SNCA-mutant cultures. Aggregation persists and progresses, ultimately resulting in the accumulation of phosphorylated α-synuclein aggregates. Impaired intracellular calcium signalling, increased basal calcium, and impairments in mitochondrial calcium handling occurred early at day 34–41 post differentiation. Once midbrain identity fully developed, at day 48–62 post differentiation, SNCA-mutant neurons exhibited mitochondrial dysfunction, oxidative stress, lysosomal swelling and increased autophagy. Ultimately these multiple cellular stresses lead to abnormal excitability, altered neuronal activity, and cell death. Our differentiation paradigm generates an efficient model for studying disease mechanisms in PD and highlights that protein misfolding to generate intraneuronal oligomers is one of the earliest critical events driving disease in human neurons, rather than a late-stage hallmark of the disease

Characterization of the Rabbit Neonatal Fc Receptor (FcRn) and Analyzing the Immunophenotype of the Transgenic Rabbits That Overexpresses FcRn

The neonatal Fc receptor (FcRn) regulates IgG and albumin homeostasis, mediates maternal IgG transport, takes an active role in phagocytosis, and delivers antigen for presentation. We have previously shown that overexpression of FcRn in transgenic mice significantly improves the humoral immune response. Because rabbits are an important source of polyclonal and monoclonal antibodies, adaptation of our FcRn overexpression technology in this species would bring significant advantages. We cloned the full length cDNA of the rabbit FcRn alpha-chain and found that it is similar to its orthologous analyzed so far. The rabbit FcRn - IgG contact residues are highly conserved, and based on this we predicted pH dependent interaction, which we confirmed by analyzing the pH dependent binding of FcRn to rabbit IgG using yolk sac lysates of rabbit fetuses by Western blot. Using immunohistochemistry, we detected strong FcRn staining in the endodermal cells of the rabbit yolk sac membrane, while the placental trophoblast cells and amnion showed no FcRn staining. Then, using BAC transgenesis we generated transgenic rabbits carrying and overexpressing a 110 kb rabbit genomic fragment encoding the FcRn. These transgenic rabbits – having one extra copy of the FcRn when hemizygous and two extra copies when homozygous - showed improved IgG protection and an augmented humoral immune response when immunized with a variety of different antigens. Our results in these transgenic rabbits demonstrate an increased immune response, similar to what we described in mice, indicating that FcRn overexpression brings significant advantages for the production of polyclonal and monoclonal antibodies