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Many factors are at play within the genome of an organism, contributing to much of the diversity and variation across the tree of life. While the genome is generally encoded by four nucleotides, A, C, T, and G, this code can be expanded. One particular mechanism that we examine in this thesis is modification of bases—more specifically, methylation of Adenine (m6A) within the GATC motif of Escherichia coli. These methylated adenines are especially important in a process called methyl-directed mismatch repair (MMR), a pathway responsible for repairing errors in the DNA sequence produced by replication. In this pathway, methylated adenines identify the parent strand and direct the repair proteins to correct the erroneous base in the daughter strand. While the primary role of methylated adenines at GATC sites is to direct the MMR pathway, this methylation has also been found to affect other processes, such as gene expression, the activity of transposable elements, and the timing of DNA replication. However, in the absence of MMR, the ability of these other processes to maintain adenine methylation and its targets is unknown.
To determine if the disruption of the MMR pathway results in the reduced conservation of methylated adenines as well as an increased tolerance for mutations that result in the loss or gain of new GATC sites, we surveyed individual clones isolated from experimentally evolving wild-type and MMR-deficient (mutL- ;conferring an 150x increase in mutation rate) populations of E. coli with whole-genome sequencing. Initial analysis revealed a lack of mutations affecting methylation sites (GATC tetranucleotides) in wild-type clones. However, the inherent low mutation rates conferred by the wild-type background render this result inconclusive, due to a lack of statistical power, and reveal a need for a more direct measure of changes in methylation status. Thus as a first step to comparative methylomics, we benchmarked four different methylation-calling pipelines on three biological replicates of the wildtype progenitor strain for our evolved populations.
While it is understood that these methylated sites play a role in the MMR pathway, it is not fully understood the full extent of their effect on the genome. Thus the goal of this thesis was to better understand the forces which maintain the genome, specifically concerning m6A within the GATC motif.
To determine if the disruption of the MMR pathway results in the reduced conservation of methylated adenines as well as an increased tolerance for mutations that result in the loss or gain of new GATC sites, we surveyed individual clones isolated from experimentally evolving wild-type and MMR-deficient (mutL- ;conferring an 150x increase in mutation rate) populations of E. coli with whole-genome sequencing. Initial analysis revealed a lack of mutations affecting methylation sites (GATC tetranucleotides) in wild-type clones. However, the inherent low mutation rates conferred by the wild-type background render this result inconclusive, due to a lack of statistical power, and reveal a need for a more direct measure of changes in methylation status. Thus as a first step to comparative methylomics, we benchmarked four different methylation-calling pipelines on three biological replicates of the wildtype progenitor strain for our evolved populations.
While it is understood that these methylated sites play a role in the MMR pathway, it is not fully understood the full extent of their effect on the genome. Thus the goal of this thesis was to better understand the forces which maintain the genome, specifically concerning m6A within the GATC motif.
ContributorsBoyer, Gwyneth (Author) / Lynch, Michael (Thesis director) / Behringer, Megan (Committee member) / Geiler-Samerotte, Kerry (Committee member) / School of Life Sciences (Contributor) / Department of Psychology (Contributor) / Barrett, The Honors College (Contributor)
Created2020-05
Description
Cancer is a disease that occurs in many and perhaps all multicellular organisms. Current research is looking at how different life history characteristics among species could influence cancer rates. Because somatic maintenance is an important component of a species' life history, we hypothesize the same ecological forces shaping the life history of a species should also determine its cancer susceptibility. By looking at varying life histories, potential evolutionary trends could be used to explain differing cancer rates. Life history theory could be an important framework for understanding cancer vulnerabilities with different trade-offs between life history traits and cancer defenses. Birds have diverse life history strategies that could explain differences in cancer suppression. Peto's paradox is the observation that cancer rates do not typically increase with body size and longevity despite an increased number of cell divisions over the animal's lifetime that ought to be carcinogenic. Here we show how Peto’s paradox is negatively correlated for cancer within the clade, Aves. That is, larger, long-lived birds get more cancer than smaller, short-lived birds (p=0.0001; r2= 0.024). Sexual dimorphism in both plumage color and size differ among Aves species. We hypothesized that this could lead to a difference in cancer rates due to the amount of time and energy sexual dimorphism takes away from somatic maintenance. We tested for an association between a variety of life history traits and cancer, including reproductive potential, growth rate, incubation, mating systems, and sexual dimorphism in both color and size. We found male birds get less cancer than female birds (9.8% vs. 11.1%, p=0.0058).
ContributorsDolan, Jordyn Nicole (Author) / Maley, Carlo (Thesis director) / Harris, Valerie (Committee member) / Boddy, Amy (Committee member) / School of Molecular Sciences (Contributor) / Department of Psychology (Contributor) / Barrett, The Honors College (Contributor)
Created2019-05
Description
Hundreds of thousands of people die annually from malaria; a protozoan of the genus Plasmodium is responsible for this mortality. The Plasmodium parasite undergoes several life stages within the mosquito vector, the transition between which require passage across the lumen of the mosquito midgut. It has been observed that in about 15% of parasites that develop ookinetes in the mosquito abdomen, sporozoites never develop in the salivary glands, indicating that passage across the midgut lumen is a significant barrier in parasite development (Gamage-Mendis et al., 1993). We aim to investigate a possible correlation between passage through the midgut lumen and drug-resistance trends in Plasmodium falciparum parasites. This study contains a total of 1024 Anopheles mosquitoes: 187 Anopheles gambiae and 837 Anopheles funestus samples collected in high malaria transmission areas of Mozambique between March and June of 2016. Sanger sequencing will be used to determine the prevalence of known resistance alleles for anti-malarial drugs: chloroquine resistance transporter (pfcrt), multidrug resistance (pfmdr1) gene, dihydropteroate synthase (pfdhps) and dihydrofolate reductase (pfdhfr). We compare prevalence of resistance between abdomen and head/thorax in order to determine whether drug resistant parasites are disproportionately hindered during their passage through the midgut lumen. A statistically significant difference between resistance alleles in the two studied body sections supports the efficacy of new anti-malarial gene surveillance strategies in areas of high malaria transmission.
ContributorsPhillips, Keeley Isabella (Author) / Huijben, Silvie (Thesis director) / Gile, Gillian (Committee member) / Young, Steven (Committee member) / School of Life Sciences (Contributor) / Barrett, The Honors College (Contributor)
Created2021-05
Description
The non-native mosquito Aedes aegypti has become a common nuisance in Maricopa county. Associated with human settlement, Ae. aegypti is known to reproduce in standing water sources both indoors and outdoors, within vessels such as tires, flowerpots, and neglected swimming pools (Jansen & Beebe, 2010). Ae. aegypti and the related Ae. albopictus are the primary vectors of the arboviral diseases chikungunya, Zika, yellow fever and dengue. Ae. aegypti tends to blood feed multiple times per gonotrophic cycle (cycle of feeding and egg laying) which, alongside a preference for human blood and close association with human habitation, contributes to an increased risk of Ae. aegypti borne virus transmission (Scott & Takken, 2012). Between 2010-2017, 153 travel-associated cases of dengue were reported in the whole of Arizona (Rivera et al., 2020); while there have been no documented locally transmitted cases of Aedes borne diseases in Maricopa county, there are no apparent reasons why local transmission can’t occur in the future via local Aedes aegypti mosquitoes infected after feeding from travelling viremic hosts. Incidents of local dengue transmission in New York (Rivera et al., 2020) and Barcelona (European Center for Disease Control [ECDC], 2019) suggest that outbreaks of Aedes borne arbovirus’ can occur in regions more temperate than the current endemic range of Aedes borne diseases. Further, while the fact that Ae. aegypti eggs have a high mortality rate when exposed to cold temperatures limits the ability for Ae aegypti to establish stable breeding populations in temperate climates (Thomas, Obermayr, Fischer, Kreyling, & Beierkuhnlein, 2012), global increases in temperature will expand the possible ranges of Ae aegypti and Aedes borne diseases.
ContributorsHon, Ruiheng (Author) / Paaijmans, Krijn (Thesis director) / Bond, Angela (Committee member) / Angilletta, Michael (Committee member) / School of Molecular Sciences (Contributor) / Barrett, The Honors College (Contributor)
Created2020-05