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Description
Glioblastoma (GB) is one of the deadliest cancers and the most common form of adult primary brain tumors. SGEF (ARHGEF26) has been previously shown to be overexpressed in GB tumors, play a role in cell invasion/migration, and increase temozolomide (TMZ) resistance.[3] It was hypothesized parental LN229 cell lines with SGEF knockdown (LN229-SGEFi) will show decreased metabolism in the MTS assay and decreased colony formation in a colony formation assay compared to parental LN229 cells after challenging the two cell lines with TMZ. For WB and co-immunoprecipitation (co-IP), parental LN229 cells with endogenous SGEF and BRCA were expected to interact and stain in the BRCA1:IP WB. LN229-SGEFi cells were expected to show very little SGEF precipitated due to shRNA targeted knockdown of SGEF. In conditions with mutations in the BRCA1 binding site (LN229-SGEFi + AdBRCAm/AdDM), SGEF expression was expected to decrease compared to parental LN229 or LN229-SGEFi cells reconstituted with WT SGEF (LN229-SGEFi + AdWT). LN229 infected with AdSGEF with a mutated nuclear localization signal (LN229-SGEFi + AdNLS12m) were expected to show BRCA and SGEF interaction since whole cell lysates were used for the co-IP. MTS data showed no significant differences in metabolism between the two cell lines at all three time points (3, 5, and 7 days). Western blot analysis was successful at imaging both SGEF and BRCA1 protein bands from whole cell lysate. The CFA showed no significant difference between cell lines after being challenged with 500uM TMZ. The co-IP immunoblot showed staining for BRCA1 and SGEF for all lysate samples, including unexpected lysates such as LN229-SGEFi, LN229-SGEFi + AdBRCAm, and LN229-SGEFi + AdDM. These results suggested either an indirect protein interaction between BRCA1 and SGEF, an additional BRCA binding site not included in the consensus, or possible detection of the translocated SGEF in knockdown cells lines since shRNA cannot enter the nucleus. Further optimization of CO-IP protocol, MTS assay, and CFA will be needed to characterize the SGEF/BRCA1 interaction and its role in cell survival.
ContributorsNabaty, Natalie Lana (Author) / Douglas, Lake (Thesis director) / Loftus, Joseph C. (Committee member) / School of Life Sciences (Contributor) / Department of Psychology (Contributor) / Barrett, The Honors College (Contributor)
Created2021-05
Description
Cancer rates vary between people, between cultures, and between tissue types, driven by clinically relevant distinctions in the risk factors that lead to different cancer types. Despite the importance of cancer location in human health, little is known about tissue-specific cancers in non-human animals. We can gain significant insight into how evolutionary history has shaped mechanisms of cancer suppression by examining how life history traits impact cancer susceptibility across species. Here, we perform multi-level analysis to test how species-level life history strategies are associated with differences in neoplasia prevalence, and apply this to mammary neoplasia within mammals. We propose that the same patterns of cancer prevalence that have been reported across species will be maintained at the tissue-specific level. We used a combination of factor analysis and phylogenetic regression on 13 life history traits across 90 mammalian species to determine the correlation between a life history trait and how it relates to mammary neoplasia prevalence. The factor analysis presented ways to calculate quantifiable underlying factors that contribute to covariance of entangled life history variables. A greater risk of mammary neoplasia was found to be correlated most significantly with shorter gestation length. With this analysis, a framework is provided for how different life history modalities can influence cancer vulnerability. Additionally, statistical methods developed for this project present a framework for future comparative oncology studies and have the potential for many diverse applications.
ContributorsFox, Morgan Shane (Author) / Maley, Carlo C. (Thesis director) / Boddy, Amy (Committee member) / Compton, Zachary (Committee member) / School of Mathematical and Statistical Sciences (Contributor) / School of Molecular Sciences (Contributor) / School of Life Sciences (Contributor) / Barrett, The Honors College (Contributor)
Created2021-05
Description
Cardiovascular disease is one of the most deadly outcomes of end stage renal disease. Bioelectrical impedance is a intriguing, yet unproven method of measuring fluid buildup in the heart, and is marketed as a early diagnostic tool for onset of cardiovascular disease. In this study, selenium supplements were given to a cohort of dialysis patients in the Phoenix metro area and their fluid tolerance was measured with thoracic biolectrical impedance. BNP was used as a correlate to see if bioelectrical impedance was correlated with heart disease. The study found no correlation between BNP and bioelectrical impedance and thus was not an accurate diagnostic tool in a medical setting.
ContributorsBrown, Patrick Michael (Author) / Johnston, Carol (Thesis director) / Orchinik, Miles (Committee member) / Tingey, Michael (Committee member) / Barrett, The Honors College (Contributor) / School of Life Sciences (Contributor) / School of Historical, Philosophical and Religious Studies (Contributor)
Created2013-05
Description
Nucleic acids encode the information required to create life, and polymerases are the gatekeepers charged with maintaining the storage and flow of this genetic information. Synthetic biologists utilize this universal property to modify organisms and other systems to create unique traits or improve the function of others. One of the many realms in synthetic biology involves the study of biopolymers that do not exist naturally, which is known as xenobiology. Although life depends on two biopolymers for genetic storage, it may be possible that alternative molecules (xenonucleic acids – XNAs), could be used in their place in either a living or non-living system. However, implementation of an XNA based system requires the development of polymerases that can encode and decode information stored in these artificial polymers. A strategy called directed evolution is used to modify or alter the function of a protein of interest, but identifying mutations that can modify polymerase function is made problematic by their size and overall complexity. To reduce the amount of sequence space that needs to be samples when attempting to identify polymerase variants, we can try to make informed decisions about which amino acid residues may have functional roles in catalysis. An analysis of Family B polymerases has shown that residues which are involved in substrate specificity are often highly conserved both at the sequence and structure level. In order to validate the hypothesis that a strong correlation exists between structural conservation and catalytic activity, we have selected and mutated residues in the 9°N polymerase using a loss of function mutagenesis strategy based on a computational analysis of several homologues from a diverse range of taxa. Improvement of these models will hopefully lead to quicker identification of loci which are ideal engineering targets.
ContributorsHaeberle, Tyler Matthew (Author) / Chaput, John (Thesis director) / Chen, Julian (Committee member) / Larsen, Andrew (Committee member) / Barrett, The Honors College (Contributor) / Department of Chemistry and Biochemistry (Contributor) / School of Life Sciences (Contributor)
Created2015-05
Description
Introduction: Human papillomavirus (HPV) infection is seen in up to 90% of cases of cervical cancer, the third leading cancer cause of death in women. Current HPV screening focuses on only two HPV types and covers roughly 75% of HPV-associated cervical cancers. A protein based assay to test for antibody biomarkers against 98 HPV antigens from both high and low risk types could provide an inexpensive and reliable method to screen for patients at risk of developing invasive cervical cancer. Methods: 98 codon optimized, commercially produced HPV genes were cloned into the pANT7_cGST vector, amplified in a bacterial host, and purified for mammalian expression using in vitro transcription/translation (IVTT) in a luminescence-based RAPID ELISA (RELISA) assay. Monoclonal antibodies were used to determine immune cross-reactivity between phylogenetically similar antigens. Lastly, several protein characteristics were examined to determine if they correlated with protein expression. Results: All genes were successfully moved into the destination vector and 86 of the 98 genes (88%) expressed protein at an adequate level. A difference was noted in expression by gene across HPV types but no correlation was found between protein size, pI, or aliphatic index and expression. Discussion: Further testing is needed to express the remaining 12 HPV genes. Once all genes have been successfully expressed and purified at high concentrations, DNA will be printed on microscope slides to create a protein microarray. This microarray will be used to screen HPV-positive patient sera for antibody biomarkers that may be indicative of cervical cancer and precancerous cervical neoplasias.
ContributorsMeshay, Ian Matthew (Author) / Anderson, Karen (Thesis director) / Magee, Mitch (Committee member) / Katchman, Benjamin (Committee member) / Barrett, The Honors College (Contributor) / School of Life Sciences (Contributor)
Created2015-05
Description
Non-small cell lung cancer (NSCLC) is the leading cause of cancer-related mortality in the USA and throughout the world. Two phenotypes that promote this deadly outcome are the invasive potential of NSCLC and the emergence of therapeutic resistance in this disease. There is an unmet clinical need to understand the mechanisms that govern NSCLC cell invasion and therapeutic resistance, and to target these phenotypes towards abating the dismal five-year survival of NSCLC. The expression of the tumor necrosis factor receptor superfamily, member 12A (TNFRSF12A; Fn14) correlates with poor patient survival and invasiveness in many tumor types including NSCLC. We hypothesize that suppression of Fn14 will inhibit NSCLC cell motility and reduce cell viability. Here we demonstrate that atorvastatin calcium treatment reduces Fn14 expression in NSCLC cell lines. Prior to Fn14 protein suppression, atorvastatin calcium modulated the expression of the Fn14 modulators P-ERK1/2 and P-NF-κβ. Atorvastatin calcium treatment inhibited the migratory capacity in H1975, H2030 and H1993 cells by at least 55%. When chemotactic migration in H2030 cells was induced by the Fn14 ligand TNF-like weak inducer of apoptosis (TWEAK) treatment, atorvastatin calcium successfully negated any stimulatory effects. Inversely, treatment of NSCLC cells with cholesterol resulted in a statistically significant increase in migration. Depletion of Fn14 expression via siRNA suppressed the migratory effect of cholesterol. Finally, atorvastatin calcium treatment sensitized cells to radiation treatment, reducing cell survival. These data suggest that atorvastatin calcium may inhibit NSCLC invasiveness through a mechanism involving Fn14, and may be a novel therapeutic target in NSCLC tumors expressing Fn14.
ContributorsCornes, Victoria Elisabeth (Author) / Stout, Valerie (Thesis director) / Whitsett, Timothy (Committee member) / Carson, Vashti (Committee member) / Barrett, The Honors College (Contributor) / School of Life Sciences (Contributor)
Created2015-05
Description
Currently in synthetic biology only the Las, Lux, and Rhl quorum sensing pathways have been adapted for broad engineering use. Quorum sensing allows a means of cell to cell communication in which a designated sender cell produces quorum sensing molecules that modify gene expression of a designated receiver cell. While useful, these three quorum sensing pathways exhibit a nontrivial level of crosstalk, hindering robust engineering and leading to unexpected effects in a given design. To address the lack of orthogonality among these three quorum sensing pathways, previous scientists have attempted to perform directed evolution on components of the quorum sensing pathway. While a powerful tool, directed evolution is limited by the subspace that is defined by the protein. For this reason, we take an evolutionary biology approach to identify new orthogonal quorum sensing networks and test these networks for cross-talk with currently-used networks. By charting characteristics of acyl homoserine lactone (AHL) molecules used across quorum sensing pathways in nature, we have identified favorable candidate pathways likely to display orthogonality. These include Aub, Bja, Bra, Cer, Esa, Las, Lux, Rhl, Rpa, and Sin, which we have begun constructing and testing. Our synthetic circuits express GFP in response to a quorum sensing molecule, allowing quantitative measurement of orthogonality between pairs. By determining orthogonal quorum sensing pairs, we hope to identify and adapt novel quorum sensing pathways for robust use in higher-order genetic circuits.
ContributorsMuller, Ryan (Author) / Haynes, Karmella (Thesis director) / Wang, Xiao (Committee member) / Barrett, The Honors College (Contributor) / School of Mathematical and Statistical Sciences (Contributor) / Department of Chemistry and Biochemistry (Contributor) / School of Life Sciences (Contributor)
Created2015-05
Description
Despite the 40-year war on cancer, very limited progress has been made in developing a cure for the disease. This failure has prompted the reevaluation of the causes and development of cancer. One resulting model, coined the atavistic model of cancer, posits that cancer is a default phenotype of the cells of multicellular organisms which arises when the cell is subjected to an unusual amount of stress. Since this default phenotype is similar across cell types and even organisms, it seems it must be an evolutionarily ancestral phenotype. We take a phylostratigraphical approach, but systematically add species divergence time data to estimate gene ages numerically and use these ages to investigate the ages of genes involved in cancer. We find that ancient disease-recessive cancer genes are significantly enriched for DNA repair and SOS activity, which seems to imply that a core component of cancer development is not the regulation of growth, but the regulation of mutation. Verification of this finding could drastically improve cancer treatment and prevention.
ContributorsOrr, Adam James (Author) / Davies, Paul (Thesis director) / Bussey, Kimberly (Committee member) / Barrett, The Honors College (Contributor) / School of Mathematical and Statistical Sciences (Contributor) / Department of Chemistry and Biochemistry (Contributor) / School of Life Sciences (Contributor)
Created2015-05
Description
Dogs' health and wellbeing is of great importance to their owners. The most common nutritional problem for pet dogs is obesity, with 22-40% of pet dogs being classified as overweight or obese. With many adverse health effects associated with obesity, this is a major concern for owners and veterinarians. The degree to which dogs enjoy consuming certain foods can have substantial implications for their body weight, so it is important to understand which aspects of foods make them appealing to dogs. This study aimed to determine whether nutritional aspects of commercial dog foods predict dogs' preferences for those foods. It was found that consumption preference is positively correlated with protein content (p < .001), therefore implying that the protein content of commercial dry dog foods may predict dogs' consumption preferences. Consumption preferences were not predicted by other available measures of food content or caloric value. Dogs' preference for foods high in protein content may be due to the satiating effect of protein. Since foods high in protein both reduce the amount of energy consumed and are found to be palatable to dogs, high-protein dog foods may offer a way for dog food manufacturers, veterinarians, and pet owners to combat obesity in pet dogs.
ContributorsPrevost, Emily Danielle (Author) / Wynne, Clive (Thesis director) / Hall, Nathaniel (Committee member) / School of Life Sciences (Contributor) / Department of Psychology (Contributor) / Barrett, The Honors College (Contributor)
Created2016-05
Description
Cancer remains one of the leading killers throughout the world. Death and disability due to lung cancer in particular accounts for one of the largest global economic burdens a disease presents. The burden on third-world countries is especially large due to the unusually large financial stress that comes from late tumor detection and expensive treatment options. Early detection using inexpensive techniques may relieve much of the burden throughout the world, not just in more developed countries. I examined the immune responses of lung cancer patients using immunosignatures – patterns of reactivity between host serum antibodies and random peptides. Immunosignatures reveal disease-specific patterns that are very reproducible. Immunosignaturing is a chip-based method that has the ability to display the antibody diversity from individual sera sample with low cost. Immunosignaturing is a medical diagnostic test that has many applications in current medical research and in diagnosis. From a previous clinical study, patients diagnosed for lung cancer were tested for their immunosignature vs. healthy non-cancer volunteers. The pattern of reactivity against the random peptides (the ‘immunosignature’) revealed common signals in cancer patients, absent from healthy controls. My study involved the search for common amino acid motifs in the cancer-specific peptides. My search through the hundreds of ‘hits’ revealed certain motifs that were repeated more times than expected by random chance. The amino acids that were the most conserved in each set include tryptophan, aspartic acid, glutamic acid, proline, alanine, serine, and lysine. The most overall conserved amino acid observed between each set was D - aspartic acid. The motifs were short (no more than 5-6 amino acids in a row), but the total number of motifs I identified was large enough to assure significance. I utilized Excel to organize the large peptide sequence libraries, then CLUSTALW to cluster similar-sequence peptides, then GLAM2 to find common themes in groups of peptides. In so doing, I found sequences that were also present in translated cancer expression libraries (RNA) that matched my motifs, suggesting that immunosignatures can find cancer-specific antigens that can be both diagnostic and potentially therapeutic.
ContributorsShiehzadegan, Shima (Author) / Johnston, Stephen (Thesis director) / Stafford, Phillip (Committee member) / School of Life Sciences (Contributor) / Barrett, The Honors College (Contributor)
Created2015-12