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- Genre: Masters Thesis
Description
Recombinant protein expression is essential to biotechnology and molecular medicine, but facile methods for obtaining significant quantities of folded and functional protein in mammalian cell culture have been lacking. Here I describe a novel 37-nucleotide in vitro selected sequence that promotes unusually high transgene expression in a vaccinia driven cytoplasmic expression system. Vectors carrying this sequence in a monocistronic reporter plasmid produce >1,000-fold more protein than equivalent vectors with conventional vaccinia promoters. Initial mechanistic studies indicate that high protein expression results from dual activity that impacts both transcription and translation. I suggest that this motif represents a powerful new tool in vaccinia-based protein expression and vaccine development technology.
ContributorsFlores, Julia Anne (Author) / Chaput, John C (Thesis advisor) / Jacobs, Bertram (Committee member) / LaBaer, Joshua (Committee member) / Arizona State University (Publisher)
Created2012
Description
The TP53 tumor suppressor gene is the most frequently mutated gene in human cancers. In the highly aggressive triple negative breast cancer (TNBC), TP53 is mutated in 80% of cases. TNBC lacks viable drug targets, resulting in a low prognosis (12.2% 5 year survivability rate). As such, the discovery of druggable targets in TNBC would be beneficial. Mutated p53 protein typically occurs as a missense mutation and often endows cancer cells with gain of function (GOF) properties by dysregulating metabolic pathways. One of these frequently dysregulated pathways is the Hippo/Yes-associated protein-1 (YAP1)/WW Domain Containing Transcription Regulator 1 (TAZ) tumor suppressor pathway. This study therefore analyzed the involvement of the Hippo/YAP1/TAZ pathway in p53-mediated breast cancer cell invasion. From an RNA-seq screen in MCF10A cell lines harboring different TP53 missense mutations, each with a differing invasive phenotype, components of the Hippo pathway were found to correlate with cell invasion. To this end, the active and inactive forms of YAP1 and TAZ were studied. Phosphorylated (inactive) YAP1 and TAZ are retained in the cytoplasm and eventually degraded. Unphosphorylated (active) YAP1 and TAZ translocate to the nucleus to activate TEAD-family transcription factors, inducing cell survival and proliferation genes leading to increased cell invasion. Using quantitative western blot analysis, it was found that inactive TAZ expression was lower in the most invasive cell lines and higher in the least invasive cell lines (p = 0.003). Moreover, the ratio of inactive TAZ protein to total TAZ protein was also shown to be predominantly lower in the invasive cell lines compared to the non-invasive lines (p = 0.04). Finally, active TAZ expression was primarily higher in p53-mutant invasive cell lines and lower in non-invasive p53 mutant cells. Additionally, although YAP1 and TAZ are thought to be functionally redundant, the pattern seen in TAZ was not seen in the YAP1 protein. Taken together, the results demonstrated here suggest that TAZ holds a more dominant role in governing TNBC cell invasion compared to YAP1 and further highlights TAZ as a potential therapeutic target in TNBC.
ContributorsGrief, Dustin (Author) / LaBaer, Joshua (Thesis advisor) / Anderson, Karen (Committee member) / Nikkhah, Mehdi (Committee member) / Arizona State University (Publisher)
Created2022
Description
Machine learning (ML) and deep learning (DL) has become an intrinsic part of multiple fields. The ability to solve complex problems makes machine learning a panacea. In the last few years, there has been an explosion of data generation, which has greatly improvised machine learning models. But this comes with a cost of high computation, which invariably increases power usage and cost of the hardware. In this thesis we explore applications of ML techniques, applied to two completely different fields - arts, media and theater and urban climate research using low-cost and low-powered edge devices. The multi-modal chatbot uses different machine learning techniques: natural language processing (NLP) and computer vision (CV) to understand inputs of the user and accordingly perform in the play and interact with the audience. This system is also equipped with other interactive hardware setups like movable LED systems, together they provide an experiential theatrical play tailored to each user. I will discuss how I used edge devices to achieve this AI system which has created a new genre in theatrical play. I will then discuss MaRTiny, which is an AI-based bio-meteorological system that calculates mean radiant temperature (MRT), which is an important parameter for urban climate research. It is also equipped with a vision system that performs different machine learning tasks like pedestrian and shade detection. The entire system costs around $200 which can potentially replace the existing setup worth $20,000. I will further discuss how I overcame the inaccuracies in MRT value caused by the system, using machine learning methods. These projects although belonging to two very different fields, are implemented using edge devices and use similar ML techniques. In this thesis I will detail out different techniques that are shared between these two projects and how they can be used in several other applications using edge devices.
ContributorsKulkarni, Karthik Kashinath (Author) / Jayasuriya, Suren (Thesis advisor) / Middel, Ariane (Thesis advisor) / Yu, Hongbin (Committee member) / Arizona State University (Publisher)
Created2021
Description
MicroRNAs (miRNAs) are 17-22 nucleotide non-coding RNAs that regulate gene expression by targeting non-complementary elements in the 3’ untranslated regions (3’UTRs) of mRNAs. miRNAs, which form complex networks of interaction that differ by tissue and developmental stage, display conservation in their function across metazoan species. Yet much remains unknown regarding their biogenesis, localization, strand selection, and their absolute abundance due to the difficulty of detecting and amplifying such small molecules. Here, I used an updated HT qPCR-based methodology to follow miRNA expression of 5p and 3p strands for all 190 C. elegans miRNAs described in miRBase throughout all six developmental stages in triplicates (total of 9,708 experiments), and studied their expression levels, tissue localization, and the rules underlying miRNA strand selection. My study validated previous findings and identified novel, conserved patterns of miRNA strand expression throughout C. elegans development, which at times correlate with previously observed developmental phenotypes. Additionally, my results highlighted novel structural principles underlying strand selection, which can be applied to higher metazoans.
Though optimized for use in C. elegans, this method can be easily adapted to other eukaryotic systems, allowing for more scalable quantitative investigation of miRNA biology and/or miRNA diagnostics.
ContributorsMeadows, Dalton Alexander (Author) / Mangone, Marco (Thesis advisor) / LaBaer, Joshua (Committee member) / Murugan, Vel (Committee member) / Wilson-Rawls, Jeanne (Committee member) / Arizona State University (Publisher)
Created2023
Description
The measurement of the radiation and convection that the human body experiences are important for ensuring safety in extreme heat conditions. The radiation from the surroundings on the human body is most often measured using globe or cylindrical radiometers. The large errors stemming from differences in internal and exterior temperatures and indirect estimation of convection can be resolved by simultaneously using three cylindrical radiometers (1 cm diameter, 9 cm height) with varying surface properties and internal heating. With three surface balances, the three unknowns (heat transfer coefficient, shortwave, and longwave radiation) can be solved for directly. As compared to integral radiation measurement technique, however, the bottom mounting using a wooden-dowel of the three-cylinder radiometers resulted in underestimated the total absorbed radiation. This first part of this thesis focuses on reducing the size of the three-cylinder radiometers and an alternative mounting that resolves the prior issues. In particular, the heat transfer coefficient in laminar wind tunnel with wind speed of 0.25 to 5 m/s is measured for six polished, heated cylinders with diameter of 1 cm and height of 1.5 to 9 cm mounted using a wooden dowel. For cylinders with height of 6 cm and above, the heat transfer coefficients are independent of the height and agree with the Hilpert correlation for infinitely long cylinder. Subsequently, a side-mounting for heated 6 cm tall cylinder with top and bottom metallic caps is developed and tested within the wind tunnel. The heat transfer coefficient is shown to be independent of the flow-side mounting and in agreement with the Hilpert correlation.
The second part of this thesis explores feasibility of employing the three-cylinder concept to measuring all air-flow parameters relevant to human convection including mean wind speed, turbulence intensity and length scale. Heated cylinders with same surface properties but varying diameters are fabricated. Uniformity of their exterior temperature, which is fundamental to the three-cylinder anemometer concept, is tested during operation using infrared camera. To provide a lab-based method to measure convection from the cylinders in turbulent flow, several designs of turbulence-generating fractal grids are laser-cut and introduced into the wind tunnel.
ContributorsGupta, Mahima (Author) / Rykaczewski, Konrad (Thesis advisor) / Pathikonda, Gokul (Thesis advisor) / Middel, Ariane (Committee member) / Arizona State University (Publisher)
Created2024
Description
During summer 2014, a study was conducted as part of the Landscape Architecture Foundation Case Study Investigation to analyze features of three sustainably designed landscapes. Each project was located in a southwest desert city: Civic Space Park in Phoenix, AZ, the Pete V. Domenici US Courthouse Sustainable Landscape Retrofit in Albuquerque, NM, and George "Doc" Cavalliere Park in Scottsdale, AZ. The principal components of each case study were performance benefits that quantified ongoing ecosystem services. Performance benefits were developed from data provided by the designers and collected by the research team. The functionality of environmental, social, and economic sustainable features was evaluated. In southwest desert cities achieving performance benefits such as microclimate cooling often come at the cost of water conservation. In each of these projects such tradeoffs were balanced by prioritizing the project goals and constraints.
During summer 2015, a study was conducted to characterize effects of tree species and shade structures on outdoor human thermal comfort under hot, arid conditions. Motivating the research was the hypothesis that tree species and shade structures will vary in their capacity to improve thermal comfort due to their respective abilities to attenuate solar radiation. Micrometeorological data was collected in full sun and under shade of six landscape tree species and park ramadas in Phoenix, AZ during pre-monsoon summer afternoons. The six landscape tree species included: Arizona ash (Fraxinus velutina Torr.), Mexican palo verde (Parkinsonia aculeata L.), Aleppo pine (Pinus halepensis Mill.), South American mesquite (Prosopis spp. L.), Texas live oak (Quercus virginiana for. fusiformis Mill.), and Chinese elm (Ulmus parvifolia Jacq.). Results showed that the tree species and ramadas were not similarly effective at improving thermal comfort, represented by physiologically equivalent temperature (PET). The difference between PET in full sun and under shade was greater under Fraxinus and Quercus than under Parkinsonia, Prosopis, and ramadas by 2.9-4.3 °C. Radiation was a significant driver of PET (p<0.0001, R2=0.69) and with the exception of ramadas, lower radiation corresponded with lower PET. Variations observed in this study suggest selecting trees or structures that attenuate the most solar radiation is a potential strategy for optimizing PET.
During summer 2015, a study was conducted to characterize effects of tree species and shade structures on outdoor human thermal comfort under hot, arid conditions. Motivating the research was the hypothesis that tree species and shade structures will vary in their capacity to improve thermal comfort due to their respective abilities to attenuate solar radiation. Micrometeorological data was collected in full sun and under shade of six landscape tree species and park ramadas in Phoenix, AZ during pre-monsoon summer afternoons. The six landscape tree species included: Arizona ash (Fraxinus velutina Torr.), Mexican palo verde (Parkinsonia aculeata L.), Aleppo pine (Pinus halepensis Mill.), South American mesquite (Prosopis spp. L.), Texas live oak (Quercus virginiana for. fusiformis Mill.), and Chinese elm (Ulmus parvifolia Jacq.). Results showed that the tree species and ramadas were not similarly effective at improving thermal comfort, represented by physiologically equivalent temperature (PET). The difference between PET in full sun and under shade was greater under Fraxinus and Quercus than under Parkinsonia, Prosopis, and ramadas by 2.9-4.3 °C. Radiation was a significant driver of PET (p<0.0001, R2=0.69) and with the exception of ramadas, lower radiation corresponded with lower PET. Variations observed in this study suggest selecting trees or structures that attenuate the most solar radiation is a potential strategy for optimizing PET.
ContributorsColter, Kaylee (Author) / Martin, Chris (Thesis advisor) / Coseo, Paul (Committee member) / Middel, Ariane (Committee member) / Arizona State University (Publisher)
Created2016
Description
Patients with malignant brain tumors have a median survival of approximately 15 months following diagnosis, regardless of currently available treatments which include surgery followed by radiation and chemotherapy. Improvement in the survival of brain cancer patients requires the design of new therapeutic modalities that take advantage of common phenotypes. One such phenotype is the metabolic dysregulation that is a hallmark of cancer cells. It has therefore been postulated that one approach to treating brain tumors may be by metabolic alteration such as that which occurs through the use of the ketogenic diet (KD). The KD is high-fat, low-carbohydrate diet that induces ketosis and has been utilized for the non-pharmacologic treatment of refractory epilepsy. It has been shown that this metabolic therapy enhances survival and potentiates standard therapy in mouse models of malignant gliomas, yet the anti-tumor mechanisms are not fully understood.
The current study reports that KetoCal® (KC; 4:1 fat:protein/carbohydrates), fed ad libitum, alters hypoxia, angiogenic, and inflammatory pathways in a mouse model of glioma. Tumors from animals maintained on KC showed reduced expression of the hypoxia marker carbonic anhydrase 9 (CA IX), a reduction in hypoxia inducible factor 1-alpha (HIF-1α) and decreased activation of nuclear factor kappa B (NF-κB). Animals maintained on KC also showed a reduction in expression of vascular endothelial growth factor receptor 2 (VEGFR2) and decreased microvasculature in their tumors. Further, peritumoral edema was significantly reduced in animals fed the KC and protein analysis showed significantly altered expression of the tight junction protein zona occludens-1 (ZO-1) and the water channeling protein aquaporin-4 (AQP4), both of which have been implicated in malignant processes in glioma, including the formation of peritumoral edema in patients. Taken together the data suggests that KC alters multiple processes involved in malignant progression of gliomas. A greater understanding of the effects of the ketogenic diet as an adjuvant therapy will allow for a more rational approach to its clinical use.
The current study reports that KetoCal® (KC; 4:1 fat:protein/carbohydrates), fed ad libitum, alters hypoxia, angiogenic, and inflammatory pathways in a mouse model of glioma. Tumors from animals maintained on KC showed reduced expression of the hypoxia marker carbonic anhydrase 9 (CA IX), a reduction in hypoxia inducible factor 1-alpha (HIF-1α) and decreased activation of nuclear factor kappa B (NF-κB). Animals maintained on KC also showed a reduction in expression of vascular endothelial growth factor receptor 2 (VEGFR2) and decreased microvasculature in their tumors. Further, peritumoral edema was significantly reduced in animals fed the KC and protein analysis showed significantly altered expression of the tight junction protein zona occludens-1 (ZO-1) and the water channeling protein aquaporin-4 (AQP4), both of which have been implicated in malignant processes in glioma, including the formation of peritumoral edema in patients. Taken together the data suggests that KC alters multiple processes involved in malignant progression of gliomas. A greater understanding of the effects of the ketogenic diet as an adjuvant therapy will allow for a more rational approach to its clinical use.
ContributorsWoolf, Eric C (Author) / Scheck, Adrienne C (Thesis advisor) / Lake, Douglas F (Committee member) / LaBaer, Joshua (Committee member) / Arizona State University (Publisher)
Created2014
Description
The rapid rate of urbanization coupled with continued population growth and anthropogenic activities has resulted in a myriad of urban climate related impacts across different cities around the world. Hot-arid cities are more vulnerable to induced urban heat effects due to the intense solar radiation during most of the year, leading to increased ambient air temperature and outdoor/indoor discomfort in Phoenix, Arizona. With the fast growth of the capital city of Arizona, the automobile-dependent planning of the city contributed negatively to the outdoor thermal comfort and to the people's daily social lives. One of the biggest challenges for hot-arid cities is to mitigate against the induced urban heat increase and improve the outdoor thermal. The objective of this study is to propose a pragmatic and useful framework that would improve the outdoor thermal comfort, by being able to evaluate and select minimally invasive urban heat mitigation strategies that could be applied to the existing urban settings in the hot-arid area of Phoenix. The study started with an evaluation of existing microclimate conditions by means of multiple field observations cross a North-South oriented urban block of buildings within Arizona State University’s Downtown campus in Phoenix. The collected data was evaluated and analyzed for a better understanding of the different local climates within the study area, then used to evaluate and partially validate a computational fluid dynamics model, ENVI-Met. Furthermore, three mitigation strategies were analyzed to the Urban Canopy Layer (UCL) level, an increase in the fraction of permeable materials in the ground surface, adding different configurations of high/low Leaf Area Density (LAD) trees, and replacing the trees configurations with fabric shading. All the strategies were compared and analyzed to determine the most impactful and effective mitigation strategies. The evaluated strategies have shown a substantial cooling effect from the High LAD trees scenarios. Also, the fabric shading strategies have shown a higher cooling effect than the Low LAD trees. Integrating the trees scenarios with the fabric shading had close cooling effect results in the High LAD trees scenarios. Finally, how to integrate these successful strategies into practical situations was addressed.
ContributorsAldakheelallah, Abdullah (Author) / Reddy, T Agami (Thesis advisor) / Middel, Ariane (Committee member) / Coseo, Paul (Committee member) / Arizona State University (Publisher)
Created2020