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Chemical Biology of Chromosome Segregation in Pseudomonas aeruginosa
No full textChromosome duplication and cell division are two critical events of a cell’s life cycle that occur with high precision across diverse environmental conditions. Inheriting a single copy of a chromosome by each daughter cell is essential for survival. An important aspect ensuring this is chromosome organization by condensins and other nucleoid-associated proteins, which, through varied levels of compaction, enables numerous DNA-dependent processes.
Passing down a fully duplicated and condensed chromosome during cell division is not a trivial job for bacteria since they efficiently adapt to the constantly changing environments and conditions that may cause DNA damage or interrupt normal progression and coordination between cell cycle events. Unlike in eukaryotes, chromosome duplication and cell division occur concurrently. To ensure the coordinated progression of the cell cycle and viability of offspring, bacteria employ a variety of pathways, some of which are known, while others are still unknown or yet to be discovered, for coordinating DNA duplication with cell division. The role of global chromosome compaction in the process of DNA duplication and cell division also remains unknown. Therefore, a better understanding of these complex processes and, more importantly, the mechanism of proteins involved in these processes could significantly enhance our existing knowledge. This, in turn, can enable us to manipulate chromosomes for various applications, find new targets for drug discovery, and even develop complementary approaches to genetic manipulations to explore cellular mechanisms of these processes in bacteria.
The current study investigated the role of two redundant systems, condensins and ParB, in the organization and assembly of replisome, segrosome, and divisome at the midcell and cell quarters in P. aeruginosa. P. aeruginosa is a Gram-negative, opportunistic human pathogen infecting immunocompromised patients and is the major cause of mortality and morbidity in patients suffering from cystic fibrosis. It has a reputation for being notoriously resistant to multiple antibiotics, making infections caused by P. aeruginosa life-threatening. Till now, the vast majority of studies have focused on multi-drug resistant aspects of P. aeruginosa, but lately, it has been gaining attention in the scientific world as an interesting model for cell cycle studies due to the discovery of new proteins and pathways associated with the cell cycle.
Previously, our lab discovered a new class of condensins, MksB, a relative of MukB, that might have comparable functions in DNA compaction and segregation in P. aeruginosa and prompted us to further explore other life cycle facets of this fascinating yet deadly pathogen. Our lab also found that the condensins (SMC and MksB) and the ParABS system, mediating dynamics, and positioning of origin of replication (oriC) are synthetically lethal. Deletion of either affected oriC dynamics, global chromosome layout, and, most importantly, disrupted chromosome segregation process as evidenced by the production of anucleate cells. These results piqued our curiosity, and further investigations revealed that the condensins and the ParABS system have additional biological roles in the biogenesis of bacterial cells.
Condensins and the ParABS system are presently thought to be involved in chromosome compaction and segregation. In the first project (Chapter 3), we investigated which proteins assembled and occupied the midcell and cell quarters. Next, we explored the sequence of events at the midcell and cell quarters during the cell cycle and proposed a mechanism for the biogenesis of new bacterial cells governed by condensins and ParB. Using fluorescence microscopy, we tracked the dynamics of key proteins involved in chromosome replication, segregation, and cell division relative to oriC. Our findings revealed that these processes are closely linked, with all three occurring at the midcell in newborn cells and later on at cell quarters.
After characterizing and establishing the role of condensins and the ParABS system in chromosome duplication and cell division process, we next characterized and employed chemical probes to study the mechanism and role of condensins and ParB in these processes. P. aeruginosa, like many other Gram-negative bacteria, poses a significant in healthcare settings due to its ability to develop antibiotic resistance. The rise in multi-drug-resistant bacteria is swiftly diminishing the effectiveness of current antibiotics, resulting in high mortality and morbidity rates, and increased public health challenges due to bacterial infections. Since most of the drugs that are currently in late-stage clinical trials are analogs of the antibiotics towards which bacteria are rapidly becoming resistant, there is a dire need to develop or screen new antibacterials that are unique and mechanistically different from the existing ones. Based on the synthetic lethality of condensins (SMC and MksB) and ParB, we developed an assay to screen a library of fungal extracts for potential inhibitors of chromosome segregation. Specifically, we screened for compounds that inhibited the ParABS system (Chapter 4) and condensins (Chapter 5) based on their increased activity in condensin and ParB mutants, respectively.
In the second project (Chapter 4), we report the discovery of a family of compounds extracted from Humicola sp. that had enhanced activity condensin mutants and were later characterized as the first known inhibitors of ParB protein. Specifically, we found new analogs of sterigmatocystin, one of which, 4-hydroxy-sterigmatocystin, 4HS, displayed an antibacterial activity and induced the phenotype typical for parAB mutants, including defects in chromosome segregation and cell division. The data describe an inhibitor of the ParAB pathway and expand the known spectrum of activities of sterigmatocystin to include bacterial chromosome segregation. Furthermore, 4HS was also found to disrupt oriC dynamics in a manner similar to ParB.
In the third project (Chapter 5), we discovered compounds derived from Aspergillus sp. that were synergistic with the deletion of parB. These compounds inhibited cell growth and induced cell lysis in P. aeruginosa. The hit compounds were found to be analogs of previously reported asteltoxins that inhibited respiration. Our subsequent experiments confirmed that our hit compound, 176LJ22M, also inhibited membrane bioenergetics by suppressing ATP levels and dissipating proton motive force (PMF), leading to growth inhibition and cell lysis. Comparison with known inhibitors of respiration further revealed that 176LJ22M not only inhibits respiration but also targets chromosome segregation, as evidenced by the increased frequency of anucleate cells.
Overall, in the current thesis, the fundamental concepts about the cell cycle and the associated proteins involved in its progression were first established. This was followed by the development of complementary approaches to genetic manipulations to explore the cellular mechanism of chromosome segregation in P. aeruginosa. Additionally, chemical probes were discovered that targeted key proteins involved in chromosome segregation, including condensins and the ParABS system. These probes enabled deeper insights into the roles of these proteins in the segregation of chromosomes during the cell cycle.
Keywords: chromosome segregation, condensins, ParABS system, cell cycle, chemical probes, sterigmatocystin, asteltoxin, Pseudomonas aeruginos
Synthesis of Folic Acid Conjugated Graphene Quantum Dot-Hexagonal Boron Nitride Nanocomposites for Targeted Cancer Cell Detection
Get PDFThe field of nanomedicine has evolved exponentially over the last decade with the development of new imagining modalities, nanoparticle types, and drug delivery systems aimed at treating and diagnosing cancer. Unfortunately, the paradigm for cancer diagnosis and treatment remains incomplete; there is a need for versatile and modular nanomaterials that act as contrast agents for the detection and characterization of pathologies. In this study, folic acid-conjugated graphene quantum dot-hexagonal boron nitride nanocomposites (FA-GQBNs) were synthesized for use as surface enhanced Raman spectroscopy (SERS) labels. Raman spectroscopy is a robust, label-free, spectroscopic technique that provides a molecular fingerprint and is commonly used for identifying unknown samples or compounds. Graphene quantum dots (GQDs) were synthesized from rice and bonded to hexagonal boron nitride (HBN) through electrostatic interactions. Next, graphene quantum dot-hexagonal boron nitride nanocomposites (GQBNs) are efficiently, inexpensively, and quickly synthesized using a bottom-up green synthesis strategy. Finally, to specifically and selectively target the folate receptor overexpressed on breast and gynecological cancer cells, folic acid (FA) was precisely conjugated to GQBNs to form FA-GQBNs. The synthesized nanocomposites were characterized using SEM, TEM, DLS, UV-Vis spectroscopy, FTIR, and Raman. The GQBNs were shown to increase cell proliferation by up to 36% when compared to the cell only group in cell viability assays while exhibiting superior optical and fluorescent properties and the FA-GQBNs were proven to be biocompatible. The FA-GQBNs were extensively characterized and their use as in vitro SERS labels as well as fluorescent labels explored. Understanding the properties and interactions of these nanocomposites provides a foundation for developing cancer therapeutics, drug delivery systems, wound-healing hydrogels, neurological treatments, and surface enhanced Raman spectroscopy nanoparticles that could be applied in the field of nanomedicine
CONSTRUCTING IDENTITY, CRAFTING CRITIQUE: MARY WOLLSTONECRAFT'S SUBVERSIVE USE OF THE TRAVELOGUE EPISTOLARY GENRE
Get PDFHistorically marginalized populations, denied access to traditional forms of political advocacy, have contributed to public discourse through their subversive uses of genres. In the field of Rhetoric and Writing, scholars have diversified their understanding of history by exploring new perspectives and voices in previously untapped genres. One such genre, the travelogue, was used by Mary Wollstonecraft as a mode of engaging in political conversations in eighteenth-century European society. Her text, Letters Written During a Short Residence in Sweden, Norway, and Denmark, serves as a unique example of the ways in which one woman utilizes the genre conventions of the travelogue to naturally discuss topics such as geopolitics, gender roles, identity, and equality. By placing herself into the role of "hero," she subverts expectations of womanhood and domesticity, demonstrating a woman’s ability to be bold, adventurous, and opinionated. At the same time, she writes this travelogue in the culturally “feminine” epistolary genre, rhetorically conforming to certain cultural gender roles in order to gain acceptance and broader readership. These letters, which are presumably written to an estranged lover, attracted readers due to their romanticism and performative intimacy. This rhetorical analysis underscores the importance of recognizing non-traditional modes of communication throughout history to enrich understanding and strive for greater inclusion of systemically overlooked contributors. Rhetorical scholars must continue to seek a radical incorporation of undiscovered genres to advance scholarship toward diversity and progress
Testing, Characterization, And Analysis Of Class F Fly Ash Geopolymers Under Oilfield Testing Conditions
Get PDFThe oil and gas industry relies heavily on Ordinary Portland Cement (OPC) for wellbore integrity,
but the limitations of OPC in extreme environments such as high temperatures and pressures have
necessitated the exploration of alternative materials. This dissertation investigates the potential of
fly ash-based geopolymers as a sustainable alternative to OPC, focusing on their unconfined
compressive strength (UCS) and compatibility with American Petroleum Institute (API) cement
testing standards. Geopolymers are an inorganic polymer resulting from the reaction of
aluminosilicate material and an alkali solution, typically comprised of potassium or sodium and
hydroxides The research begins with an overview of the environmental impact of traditional
cement production and the necessity for robust wellbore materials. It details the standard testing
methods for OPC, including preparation, curing, and mechanical testing procedures, and extends
these methods to geopolymers. The effects of various curing conditions and chemical compositions
on the UCS of geopolymers are evaluated through a series of experiments. Results demonstrate
significant variability in geopolymer performance, which is attributed to the inherent differences
in fly ash composition and the lack of standardized testing protocols. Enhanced investigation
techniques, such as Nuclear Magnetic Resonance (NMR) and Scanning Electron Microscopy
(SEM), provide deeper insights into the microstructural properties of geopolymers and their
correlation with mechanical performance. A comparative analysis of geopolymers and traditional
API cements under similar conditions reveals that while geopolymers offer potential benefits, their
inconsistent performance poses challenges for widespread adoption. The dissertation advocates
for the development of standardized testing procedures specific to geopolymers to ensure reliable
performance metrics. It explores the implications of using geopolymers in geothermal well
completions, suggesting that with further research and standardization, geopolymers could become
a viable alternative in high-temperature applications. The study concludes with a summary of vii
findings and recommendations for future research, emphasizing the need for a comprehensive
understanding of geopolymer chemistry and the establishment of industry-wide standards to
facilitate their adoption in oil and gas operations
The Role of Moral Foundations in the Energy Transition
Get PDFPublic support for renewable energy and climate policies is crucial for the success of the energy transition. While previous research has established the influence of political ideology on energy preferences, the role of moral foundations in shaping these attitudes remains underexplored. This study examined the relationships between moral foundations and support for renewable energy (solar and wind) and fossil fuel reduction policies, while controlling for political orientation and other relevant factors.
Using data from an online national survey of 2,188 U.S. adults, we employ binary logistic regression analyses to test the independent associations of the five moral foundations (care, fairness, authority, loyalty, purity) with energy policy preferences. The results reveal that the individualizing foundations of care and fairness consistently predict higher support for renewable energy and lower support for fossil fuels, even after accounting for political ideology. In contrast, the binding foundation of authority is negatively associated with renewable energy support, while loyalty predicts opposition to decreasing fossil fuel use. Results also demonstrate that moral foundations play a distinct role in shaping public attitudes towards energy transition policies, beyond the influence of political orientation. The care and fairness foundations emerge as potential drivers of support for renewable energy and climate action across the political spectrum. Conversely, the authority and loyalty foundations may contribute to resistance to energy system changes, particularly among conservatives.
We highlight the importance of considering moral concerns in the design and communication of energy transition strategies. Policymakers and advocates could leverage moral foundations theory to craft targeted messages and policies that resonate with different segments of the public. By aligning climate and energy initiatives with widely shared moral values, it may be possible to build broader coalitions of support for the energy transition. Further research is needed to test the effectiveness of morally-framed interventions in shifting energy attitudes and behaviors.N
THE IMPACT OF ADDITIVES IN ACIDIC ZEOLITES ON HYDROCARBON-BASED REACTIONS
No full textThe incorporation of metal additives into zeolites has allowed a much higher degree of zeolite tunability, including altering the original active sites, introducing new active sites, modifying pore structures, and introducing bifunctional properties. As a result, these modifications have a profound impact on the catalyst performance, altering product selectivity, catalyst activity, and catalyst stability. This thesis investigates the role of several metal additives, including sodium, calcium, cobalt and lanthanum, focusing on their impact during isooctane cracking and aromatic alkylation reaction between toluene and isopropanol.
The first part of this thesis explores isooctane cracking over faujasite zeolite as probe enabling the quantification of protolytic cracking, dehydrogenation, and hydrogen transfer pathway. The development of this probe was possible by the addition of metal cations such as Na, Ca, Co as additives to modify the rates of these reaction pathways. This has enabled a quantitative assessment of cation titration at not only promoting new pathways, but selectively titrating sites that are most active for protolytic cracking. We further contrast this simple approach in ternary diagrams that visually depict the contributions of catalyst modifications and reaction conditions on the three parallel reactions, revealing that Na selectively titrates sites responsible for protolytic cracking while Co promotes dehydrogenation reactions. Finally, we reveal that the incorporation of La, often added to Y zeolites in industrial catalytic cracking operations, enhances both hydrogen transfer rates and dehydrogenation rate of alkanes.
The second and third part of the thesis primarily focuses on La as an additive. The second part is a continuation of La modified Y zeolite from the first part. This section focuses the impact of La introduction as an additive via ion exchange on the physical and chemical properties of low Si/Al Y zeolite, including the preservation of zeolitic structural integrity and acid sites modifications. The addition of La has a pronounced effect on the isooctane cracking performance of zeolite Y, altering the reaction pathway selectivity and cracking activity.
Finally, the final part of the thesis investigates the effects of La incorporation in H-MOR via wetness impregnation and focuses on the impact of La on shape selectivity and catalysts' stability against coking during the alkylation between toluene and isopropanol. The addition of La alters the pore size of the MOR zeolite via pore constriction. Furthermore, La also enhances the hydrogen transfer activity between carbenium ion and coke deposited. This modification leads to improved product selectivity and catalyst stability, highlighting the potential of La-modified zeolites in enhancing catalytic performance.
Collectively, these findings deepen our understanding of the role of these additives in zeolite catalysis, offering insights for optimizing catalyst synthesis for hydrocarbon reactions
CONSTRAINING INITIATION OF RIFTING IN A MAGMA POOR SYSTEM: LAKE TANGANYIKA
No full textThe Lake Tanganyika rift is commonly cited as a modern analog for early stages of continental rifting. It lies within the western branch of the East African Rift system (EARS). The modern Tanganyika rift consists of distinct normal faults that alternate facing direction down the rift axis with footwall relief of 2.5 km. Adjacent to faults, hanging wall subsidence has produced sub-basins with > 5 km of sediment fill. The age of rift initiation and uplift and subsidence along Lake Tanganyika is poorly constrained, however, and has mostly been inferred from the basin-fill, but these estimates are highly dependent upon assumptions of long-term rates of erosion and sediment accumulation. The lack of temporal constraints on the initiation and spatial variability of rifting within the Tanganyika rift has led to differing models posed for its formation: 1) the extensional uplift and associated subsidence initiated in the central part of the present rift and then propagated north and south over time, or 2) the extension initiated roughly coevally along the rift axis but differed in magnitude of extension along the rift axis. Testing these models requires better constraints on magnitude and timing of footwall uplifts along the rift. Low-temperature thermochronology can capture tectonic/erosional events by tracking the exhumation of minerals within rocks from depth following normal faulting through fission track analysis. Previous studies have applied low-temperature thermochronology in other rift segments of the western branch, but no studies have included the footwall uplifts of Lake Tanganyika. Here, I apply apatite fission-track (AFT) analyses to detrital river sand samples collected along two footwall uplifts along the Tanganyika rift axis to track rock cooling and to infer the associated exhumation history. Apatite fission-track analysis of these detrital samples yields a broad range of single-grain ages from Miocene (13 Ma) to Silurian (366 Ma) in central Lake Tanganyika and Eocene (41 Ma) to Neoproterozoic (842 Ma) in southern Lake Tanganyika. Watershed analysis of the sampled river in the central region exhibits a mature geometry in the central basin while the southern exhibits a younger morphology, with sediment sources occurring at higher elevations. The cooling age populations in central Lake Tanganyika are consistent with previous basement apatite fission-track ages from the Lake Albertine Rift to the north and Lake Malawi Rift to the south, all which are interpreted to result from Cenozoic rifting along rift flanks in the western branch. However, the southern sample captures cooling associated with Permo-Triassic to Cretaceous rifting. These AFT data suggest that rifting nucleated in the center basins then continued along the rift axis as opposed to coeval rift initiation along pre-existing shear zones across all subbasins in Lake Tanganyika
POSTSECONDARY TRANSITION OF INDIGENOUS STUDENTS WITH DISABILITIES
Get PDFIndigenous students with disabilities (ISWD) are proportionally the most represented racial demographic receiving special education services under IDEA (2004), which mandates postsecondary transition services for students with an IEP. Despite their disproportionate representation, ISWD are almost entirely absent from postsecondary transition research, resulting in a lack of evidence-based practices to support their transitions. This dissertation aims to identify evidence-based strategies to make transition planning culturally responsive for ISWD, proposing a model for braiding evidence-based practices. Chapter 2 reviews the literature on evidence-based practices and predictors, revealing that ISWD constitute less than 1% of the samples used to identify and classify these practices. In Chapter 3, a panel of Indigenous knowledge holders shares their expertise on the relevance of transition assessments for ISWD, identifying five key areas for teachers to consider: culture, first-generation students, guidance/mentorship, family involvement, and financial literacy. Chapter 4 presents a phenomenological study in which family members of ISWD discuss their experiences with transition planning and suggest improvements, including awareness of resources, employment support, and financial literacy education. Chapter 5 synthesizes the findings from the previous chapters, weaving them together to propose strategies that support ISWD in the transition process. The dissertation concludes with a discussion of areas for future research and implications for practitioners
WHEN TO FOLLOW, WHEN TO BREAK, WHO TO BLAME: CITIZEN PERCEPTIONS OF STREET-LEVEL SERVICE INTERACTIONS
Get PDFIn this dissertation, I combine insights from public administration with those of political science, public policy, and social psychology to better understand how citizens think about and understand their governments. In each of three studies, I unpack a different decision-making process with relevance to public administration. The first explores how information about clients shapes citizens’ agreement with street-level bureaucrats’ rule compliance (or lack thereof). The second is similar, this time assessing how client information influences how citizens assign blame when clients experience negative outcomes. The third focuses on factors that drive bureaucrats to support other bureaucrats’ rule compliance decisions, this time paying special attention to the effects of respondents’ just-world beliefs.
While I expected that clients’ identities—and particularly their identity congruence with respondents—would significantly affect each of these processes, what I find is more interesting and complex. Though they did not express strong disagreement with bureaucrats’ prosocial rule-breaking decisions—or decisions to break the rules for the explicit purpose of better serving a client— citizens always preferred that bureaucrats followed agency rules, regardless of the client’s identity, deservingness, and outcome. When assigning blame for a client’s negative outcome, citizens overwhelmingly blamed the client, allocating over twice as much blame to the client than the serving bureaucrat or agency. While, this time, the client’s deservingness had large effects on citizens’ decisions, even deserving clients received more blame than any other category. Finally, respondents—both citizens and bureaucrats—with high just-world beliefs were much more likely to support bureaucrats’ rule decisions, irrespective of the actual decision. As I will argue, together, the findings provide room for optimism in some ways while painting a worrying picture for social equity in others
Experiences of Social Isolation in Rural Communities: A Multidisciplinary Approach
Get PDFRural communities often struggle due to a lack of funding, resources, and support, which can put residents at risk for adverse experiences. Merely living within a rural area is considered an influential contributing risk factor for experiencing social isolation and loneliness. Experiences of social isolation and loneliness are classified as major public health concerns due to the damaging effects they can have on mental and physical health. However, little is known about neural responses to social isolation seen in rural residents. Thus, this study aimed to examine the issues rural communities in Oklahoma face, how these communities respond to experiences of social isolation, and how different experiences and behavioral traits affect this response. Electroencephalography (EEG) was used to assess differences in neural activity in response to rejection; while undergoing EEG, healthy adults across rural Oklahoma played a cyberball game specifically designed to elicit acute feelings of social rejection. In addition, a detailed survey was distributed that encompasses many facets of social experience that may influence their response to social isolation. Specifically, adverse childhood experiences (ACEs), resilience, and social support may serve as important factors for explaining differences in these responses. Results suggest that social support, resilience, and childhood trauma play important roles in how an individual responds to rejection. Social support appeared to foster the formation and continuation of a resilient mindset. Resilient and non-resilient individuals responded differently to rejection, further indicating the importance of a resilient mindset. Understanding the interactions between these variables and an individual’s neural response to rejection will inform future studies on rural communities, which may provide more opportunities for improvement within these communities. The use of a multidisciplinary approach allows for a thorough investigation of adverse childhood experiences, resilience, and social support in the context of rural communities