Now showing items 21-40 of 1675

    • Molecular Mechanisms Underlying the Metastasis Suppressor Activity of NME1 in Malignant Melanoma

      Pamidimukkala, Nidhi Vela; Kaetzel, David M. (2019)
      Metastatic melanoma is exceedingly lethal and is responsible for the majority of skin cancer related deaths. Impactful research on melanoma metastasis is crucial to improving prognosis. The discovery of metastasis suppressor genes, genes that inhibit metastasis but do not affect tumor growth, have advanced the field of metastasis research. NME1 (also referred to as NDPK-A or NM23-H1) was the first identified metastasis suppressor gene. NME1 is a multifaceted molecule, executing numerous cellular functions which are attributable to the suppressor phenotype. Our lab previously identified a signature of NME1-regualted genes which have prognostic value in human melanoma. As such, we hypothesize that NME1 possess an understudied transcriptional activity to regulate one or more of these signature genes. In the present study, we identified Aldolase C (ALDOC) from the gene signature as an ideal candidate to assess NME1 transcriptional activity. NME1 induced mRNA and protein expression of ALDOC in multiple melanoma cell lines. Transcriptional regulation was evidenced by elevated expression of ALDOC pre-mRNA and activation of the ALDOC promoter in NME1-expressing melanoma cells. Employing chromatin immunoprecipitation, we demonstrated NME1 localizes to the ALDOC gene and enriches the presence of active transcription indicators at the ALDOC promoter. Together, we provide novel systematic evidence for NME1 transcriptional activity, which may further be explored as a mechanism for the metastasis suppressor function. Furthermore, we establish unequivocal metastasis suppressor capability of NME1 using a transgenic mouse model susceptible to developing melanoma upon exposure to ultraviolet (UV) radiation. Mice deficient in NME1 expression experienced significantly increased melanoma metastases to the lung and lymph nodes compared to wild-type mice. Taken together, our studies demonstrate NME1 as a robust metastasis suppressor of UV-induced melanoma, and provide evidence for novel transcription factor activity of NME1. We aim to enhance our understanding of melanoma metastasis through our continued research of the metastasis suppressor gene, NME1.
    • Regulation of CD8+ T cell differentiation by cytokines and T-box transcription factors

      Reiser, John Wyatt; Singh, Nevil (2019)
      A protective immune response requires naïve CD8+ T cells to differentiate into effector cells, some of which persist as a memory population to protect against future threats. This differentiation program is controlled by a transcriptional network involving the T-box transcription factors T-bet and Eomesodermin (Eomes). These factors are in turn differentially regulated by cytokines in the local milieu. The precise architecture of this regulatory network in terms of the individual, synergistic and redundant roles of T-bet, Eomes and critical cytokines is still poorly understood. Here we sought to clarify these regulatory pathways, using mice deficient in T-bet, Eomes or both. Our studies identify a key role for Eomes, but not T-bet, in regulating IL-10 expression by CD8+ T cells early after activation. We further reveal a feedback loop where Eomes and IL-10 cooperatively enhance the expression of the lymph-node homing selectin CD62L. Consistent with the importance of CD62L in promoting memory T cell homing to secondary lymphoid organs (SLOs), we identify a role for CD8+ T cell-intrinsic IL-10 in promoting the long-term persistence of memory cells in vivo. In contrast, T cells that home away from the SLOs become terminal effector T cells. Using tumor and infectious disease models, we report that these cells experience a second wave of T-bet induction in the tissue, which may contribute to a heightened effector response. We also define distinct roles for T-bet and Eomes in regulating effector and memory genes, highlighting their redundant role in repressing Tc2 and Tc17 differentiation in CD8+ T cells. Finally, we demonstrate that in the context of a tumor-specific immune response, the absence of both T-box transcription factors severely limits tumor infiltration without significant alteration in early proliferation or effector functions. Taken together, these findings offer new insights into how T-bet, Eomes and IL-10 regulate effector and memory responses in distinct tissues during the immune response. Future developments targeting individual aspects of T-box and cytokine signaling in T cells may help engineer precise outcomes in the context of tumor immunotherapies, vaccination and transplantation.
    • Pseudomonas aeruginosa adaptation, pathogenicity, and persistence in the environment and cystic fibrosis airway.

      Chandler, Courtney; Ernst, Robert K.; 0000-0003-2076-3665 (2019)
      Pseudomonas aeruginosa is an opportunistic Gram-negative bacterium associated with airway infections of patients with cystic fibrosis (CF). Lipid A is the membrane anchor of lipopolysaccharide, the dominant component of the outer leaflet of the outer membrane of Gram-negative bacteria. Lipid A structure can be modified via a number of biosynthetic enzymes. Here, we describe two -hydroxylase enzymes, LpxO1 and LpxO2, capable of catalyzing 2-hydroxylation of lipid A in P. aeruginosa. We additionally characterize their role in persistence and infectivity. Phylogenetic analysis suggests at least one lpxO originated from lateral gene transfer and the gene duplication is a recurring feature in Pseudomonas evolution. To determine the roles of LpxO1/2 in vivo, we used a rapid extraction from lavage fluid to describe the structure of P. aeruginosa lipid A isolated from the lungs of mice after intranasal infection. Lipid A is also known to be altered during adaptation to the lungs of CF patients, where P. aeruginosa can colonize and cause infection throughout a patient’s lifetime. This is a leading contributor of morbidity and mortality in this patient population, and thus the genetic and phenotypic adaptations that P. aeruginosa undergoes during CF airway infection are of interest. We used whole-genome sequencing of 130 P. aeruginosa isolates, including 81 from young children with CF, to define early-stage genetic adaptation events, including lipid A modification. We additionally investigate the influence of patient region of residence on such adaptation. With these data, we provide the first longitudinal analysis of P. aeruginosa genetics in young patients with CF in the United States. We additionally analyzed ten sublines of laboratory-adapted strain PAO1 to determine the level of microevolution experienced by this set of isolates. In total, our analyses contribute to our understanding of lipid A structure, synthesis, and modification in P. aeruginosa, and our sequencing data will serve as a resource for the entire CF and Pseudomonas community.
    • The Role of Testisin and PAR-2 Signaling in Ovarian Cancer Metastasis

      Conway, Gregory David; Antalis, Toni M. (2019)
      Ovarian cancer is the leading cause of death among gynecological cancers in the United States. Ovarian cancer employs a unique mode of metastasis, as tumor cells disseminate within the peritoneal cavity, colonizing in several sites and driving the accumulation of ascites. Tumor recurrence and metastasis are significant factors contributing to poor prognosis. The membrane-anchored serine protease testisin is aberrantly expressed in ovarian tumors and it’s only known substrate, protease activated receptor-2 (PAR-2), is also overexpressed in ovarian tumors. In this study I have examined 1) the role of testisin and PAR-2 signaling in ovarian tumor metastasis and 2) determined whether testisin and other membrane-anchored serine proteases may be anti-cancer therapeutic targets. We generated human ovarian ES-2 tumor lines that express testisin or the catalytically inactive testisin mutant S238A and explored the role of constitutive testisin expression in late stage ovarian cancer. We show that testisin stimulates the activation and internalization of PAR-2 resulting in decreased expression of ANG2 and ANGPTL4. Using a preclinical xenograft model of late stage ovarian cancer, we find that testisin activity in ovarian cancer cells reduces intra-peritoneal tumor dissemination, tumor burden and ascites formation. Analyses of the tumors showed that testisin activity downregulates the expression of ANG2 and ANGPTL4 in vivo as well as in vitro. To examine membrane-anchored serine proteases as therapeutic targets in cancer, we utilized an engineered anthrax toxin pro-drug strategy requiring proteolytic cleavage of a protective antigen (PrAg) for activation. We explored the efficacy of these engineered PrAg toxins in several intraperitoneal models of ovarian cancer metastasis and show PrAg treatment reduced tumor burden in both an intraperitoneal NCI/ADR-Res xenograft and an ES-2 minimal residual disease model, which replicates debulking surgery in ovarian cancer patients, but had no effect in a syngeneic ID-8-Luc xenograft model of ovarian cancer. Additionally, we found that engineered PrAg toxins are capable of inducing lung and pancreatic cancer cellular death in vitro. Data presented herein provide new insight into the potential role of testisin and PAR-2 signaling in ovarian cancer metastasis and suggests membrane-anchored serine proteases as a potential therapeutic target for metastatic ovarian cancer.
    • Characterization of PfCRT F145I in piperaquine-resistant Plasmodium falciparum isolates from Cambodia through zinc-finger nuclease-mediated gene editing

      Shrestha, Biraj; Takala-Harrison, Shannon (2019)
      ACTs are the first-line treatment for clinical malaria in the malaria-endemic world and have reduced malaria-associated mortality and morbidity. However, the recent emergence of Plasmodium falciparum that is resistant to both the artemisinins and key partner drugs, piperaquine, in Cambodia and nearby countries in GMS poses a threat to the control and elimination of malaria. Identification and validation of molecular markers of antimalarial drug resistance provide surveillance tools to monitor resistance and inform drug policy decisions and insights into the molecular mechanisms underlying resistance. Previous studies have found that F145I mutation within the PfCRT and plasmepsin2/3 gene copy number are associated with resistance to piperaquine. When PfCRT F145I is introduced into Dd2 of P. falciparum, it confers piperaquine resistance. In this study, we will use gene-editing approaches to remove F145I from field isolates that contain both this mutation and amplified plasmepsin2/3, to quantify the effect on malaria parasite susceptibility to piperaquine.
    • XIAP-p47 Pairing Activates the Immune Deficiency Pathway in the Lyme Disease Tick Ixodes scapularis

      McClure Carroll, Erin; Pedra, Joao H. F. (2019)
      Globally, vector-borne diseases account for 17% of all infectious diseases. Most vectors are blood-feeding arthropods, which transmit bacterial, viral, and parasitic diseases to humans and animals. The tick Ixodes scapularis transmits seven pathogens, including Borrelia burgdorferi, the agent of Lyme disease. Lyme disease is the most important vector-borne disease in the United States and causes an estimated 329,000 infections annually. Best described in the model organism Drosophila melanogaster, the arthropod immune deficiency (IMD) pathway responds to microbial infection through activation of Relish, a nuclear factor (NF)-κB family transcription factor. In I. scapularis ticks, the E3 ubiquitin ligase X-linked inhibitor of apoptosis (XIAP) regulates the IMD pathway through ubiquitylation. Yet, the tick genome notably lacks homologs to genes encoding key IMD pathway proteins as described in Drosophila. How XIAP activates the IMD pathway in response to microbial infection is poorly characterized and targets of XIAP-mediated ubiquitylation remain unknown. In this study, we identified the XIAP enzymatic substrate p47 as a positive regulator of the I. scapularis IMD network. XIAP polyubiquitylates p47 in a lysine (K)63-dependent manner and interacts with the ubiquitin-like (UBX) domain of p47. p47 also binds to Kenny (IKKγ/NF-κB essential modulator [NEMO]), the regulatory subunit of the inhibitor of NF-κB kinase (IKK) complex. Replacement of the amino acid lysine with arginine in the p47 linker region completely abrogated molecular interactions with Kenny. Furthermore, reduction of p47 transcription levels through RNA interference in I. scapularis limited Kenny accumulation, reduced phosphorylation of IKKβ (IRD5), and impaired cleavage of the NF-κB molecule Relish. Accordingly, disruption of p47 expression increased microbial colonization of the tick-transmitted spirochete B. burgdorferi and the rickettsial agent Anaplasma phagocytophilum. In summary, we demonstrated that XIAP ubiquitylates p47 in a K63-dependent manner, culminating in Relish activation and antimicrobial responses. Manipulating immune signaling cascades in I. scapularis may lead to innovative approaches to reducing the burden of tick-borne diseases.
    • Characterizing Genetic Drift and Migration between Populations

      Harris, Daniel Nathan; O'Connor, Timothy D. (2019)
      Genetic drift and migration are two of the main mechanisms of evolution at the genetic level. These mechanisms greatly impact the genetic relatedness of populations. Therefore, by studying how genetic drift and migration impact populations, it is possible to reconstruct elements of population history throughout a geographic region. This thesis examines how genetic drift and migration rapidly impact human population structure. My first aim studies the peopling of the Peruvian region and models migration patterns. Here, I suggest that the peopling of Peru was a rapid process that occurred by 12,000 years ago. Furthermore, migration modeling suggests that the majority of migration in Peru is from populations in the high altitude Andes Mountains to the low altitude coast and Amazon. My second aim, then analyzes genetic drift and migration on the fatty acid desaturase gene region, where there is a suggestion of a public health problem due to the apparent fixation of the low efficient ancestral haplogroup in Native Americans. I confirm the near fixation of the ancestral haplogroup in Native American ancestry, and suggest that understanding the frequency of the ancestral haplogroup in the Native American founding population is crucial for determining how this apparently detrimental haplogroup reached near fixation. Furthermore, I suggest that the global distribution of the haplogroup is not indicative of Neanderthal introgression and is instead likely an ancient polymorphism that arose during the divergence of modern humans and Neanderthals. For my final aim, I use rare variant and haplotype based approaches to discern the population structure and evolutionary history of Samoans. I suggest that the early population size was small and a strong population expansion occurred approximately 35 generations ago, consistent with the Samoan archaeological record. In addition, Samoan population structure is likely strongly driven by urbanization, and not biogeography. In sum, this thesis demonstrates that migration and genetic drift can rapidly impact the genetic variation in populations. In addition, I demonstrate that rare variant and haplotype based approaches are essential for examining the impact of migration and genetic drift between populations within an extremely recently founded region with minimal geographic barriers.
    • From Cell Fate to Terminal Differentiation – Insights into Inner Ear Hair Cell Development from Cell Type-Specific Transcriptomic Analyses

      Matern, Maggie S.; Hertzano, Ronna P. (2019)
      Hair cells (HCs), which are specialized neuroepithelial sensory cells of the inner ear, are responsible for conveying information about the environment, such as sound and movement, to the brain. In adult mammals, HCs do not spontaneously regenerate. Therefore, damage that occurs to HCs after noise or ototoxic drug exposure often leads to a permanent loss of inner ear function. It is estimated that 1/3 of individuals over the age of 65 experience hearing loss that could benefit from use of a hearing aid, representing a significant healthcare burden in the United States. It is for these reasons that the NIH National Institute on Deafness and Other Communication Disorders has set understanding the normal development of the auditory and vestibular systems as the Hearing and Balance Research Priority Area 1 of their 2017-2021 Strategic Plan, so that regenerative therapies can be better formulated. Here I have utilized cell type-specific ribosomal immunoprecipitation analyses to study HC gene expression, with the goal of furthering our understanding of their normal development. First, research into control of inner ear development has indicated that early HC differentiation appears to be dependent on the expression and normal functioning of the transcription factors ATOH1, POU4F3, and GFI1. While Pou4f3 and Gfi1 expression seems to be downstream of ATOH1, and the expression of Gfi1 is dependent on POU4F3, the signaling cascades downstream of GFI1 and its role in HC development are completely unknown. Here I have identified a critical role for GFI1 in HC maturation, namely through repression of a neuronal transcriptional program that is normally present in cochlear HCs during early embryonic development. Later in this developmental program, cochlear HC precursors must further differentiate into either inner or outer HCs, however, apart from the recently described INSM1 in embryonic development, no transcription factors have been implicated in the differentiation of outer HCs. It was therefore my second goal to identify and validate candidate downstream targets of helios, a newly identified postnatal outer HC transcription factor. Finally, I have developed and validated a model for ribosome immunoprecipitation and translatome analysis to study zebrafish HC gene expression – the Tg(myo6b:RiboTag) zebrafish.
    • Functional Status and Muscle Evaluation in Older Women with Urinary Incontinence.

      Sanses, Tatiana Vyacheslavovna Deniseiko; Ryan, Alice (2019)
      Background: Urinary incontinence (UI) and mobility limitations are prevalent in older women suggesting common pathophysiology. Objectives: 1) to evaluate Short Physical and Performance Battery (SPPB) and Modified Physical Performance Test (MPPT); 2) to describe lower extremities/pelvic floor muscles among older women with UI. Methods: Women age ≥ 70 years with UI underwent functional testing, clinical, and magnetic resonance imaging evaluation. Spearman correlation coefficients were calculated between UI and mobility limitations. Results: MPPT was inversely moderately associated with UI severity impact on daily activities. SPPB was negatively associated with UI but findings were not statistically significant. Levator ani muscles were atrophic in all participants. Obturator and gluteal muscles had mild and moderate fatty infiltration, respectively. Conclusions: Lower MPPT is associated with increased UI severity impact on daily activities. Further research into shared pathophysiology between UI and mobility
    • Molecular Determinants of Pulmonary Mucormycosis and Aspergillosis

      Watkins, Tonya Nicole; Bruno, Vincent, Ph.D. (2019)
      Mucormycosis and aspergillosis are invasive fungal infections with very limited treatment options and extremely high mortality rates. During both pulmonary mucormycosis and aspergillosis, inhaled fungal spores must adhere to and invade airway epithelial cells in order to establish infection. The molecular mechanisms governing these interactions in the context of each disease are not completely understood. Mucormycoses are caused by fungi belonging to the Order Mucorales, with ~70% of all cases caused by Rhizopus species. To better understand the molecular mechanisms of fungal invasion during mucormycosis, an unbiased survey of the host transcriptional response to Mucorales infection in in vitro and in vivo murine models of pulmonary mucormycosis using RNA-seq was performed. Network analysis revealed activation of host receptor tyrosine kinase (RTK) signaling pathways and progesterone (PG) signaling pathways. By combining established models of mucormycosis, transcriptomics, cell biology, and pharmacological approaches, we demonstrated that Mucorales activate epidermal growth factor receptor (EGFR), erb-b2 tyrosine kinase 2 (ErbB2), and platelet-derived growth factor receptor (PDGFR) signaling to induce fungal uptake into airway epithelial cells. Furthermore, we demonstrated that inhibition of EGFR signaling with existing FDA-approved cancer drugs significantly increased survival following Rhizopus delemar infection in mice. We also demonstrated that inhibition of progesterone receptor (PGR) signaling decreases R. delemar invasion of airway epithelial cells in vitro. These studies enhance our understanding of how Mucorales invade host cells during the establishment of pulmonary mucormycosis and provide a proof-of concept for repurposing FDA-approved cancer drugs that target RTK function. Aspergillus fumigatus is responsible for 90% of all aspergillosis cases. To better understand how A. fumigatus senses and responds to airway epithelial cells during pulmonary aspergillosis, we used RNA-seq to analyze the transcriptomes of two commonly used clinical A. fumigatus isolates, Af293 and CEA10, during an in vitro infection model of airway epithelial cells. In this analysis, we identified 47 genes that were up-regulated in both strains and enriched for genes involved in iron acquisition, a mechanism required for A. fumigatus virulence in the mammalian host. Knowledge gained from this work could aid in the identification of therapeutic and prevention targets to combat mucormycosis and aspergillosis.
    • Association between Iron Deficiency with or without Anemia and Infants’ Cognitive, Motor, and Socio-emotional Development: Cross-sectional and Longitudinal analyses

      Yimgang Ngandjouong, Doris P.; Black, Maureen M.; 0000-0002-3909-4839 (2019)
      Background: Early child development is a major public health concern worldwide as it has long-term health and economic effects that extend beyond childhood. Iron deficiency (ID) is the most common micronutrient deficiency worldwide with the highest prevalence in low- and middle-income countries. ID may impact child development early in life. Objectives of this study were to investigate the relationship between ID and child development and to identify moderators of this association among infants. Methods: Secondary data analyses of a nutritional and early learning controlled trial were performed. Participants: 513 children age 6-14 months from rural India were enrolled and followed for 12 months. Measures: At enrollment, 6-month and 12-month follow-up, child development was assessed using Mullen Scales for Early Learning (visual reception, fine/gross motor and receptive/expressive language domains) and Behavior Observation Inventory of the Bayley Scales of Infant and Toddler Development (socio-emotional domain). Analysis included linear mixed effects models accounting for village-level clustering (all aims) and repeated measures (Aims 2 and 3.1). To assess moderating effects of child age and sex, maternal education, household assets, and home environment, interaction terms were added in models. Results: At enrollment, 47% of infants were girls, 72% had ID, 66% had anemia, and the mean age was 8.6 months. Mean maternal age was 23 years and 43% of mothers were anemic. Adjusted analyses found weak to no evidence of associations between ID and child development at enrollment or over a 12-month period. Child age and sex, maternal education, household assets, and home environment did not moderate associations of interest. Conclusion and future directions: In this rural sample, ID was not associated with child development using observational study designs. In order to properly assess the onset and duration of ID and its associations with child development, further studies need to enroll pregnant women and follow up newborns during their first years of life. It is necessary to assess iron measures and child development at multiple time points throughout the study period. Implications: Because of the pervasive nature of ID in this population, there is a need for nutritional interventions to reduce micronutrient deficiencies.
    • Role of Vaccine-Induced IgG in Protection Against Bordetella Pertussis

      Masterson, Mary; Pasetti, Marcela F. (2019)
      Bordetella pertussis is a highly infectious respiratory pathogen that can induce severe bronchopneumonia and respiratory failure in infants (whooping cough). Vaccine formulations consisting of Diphtheria toxoid, Tetanus toxoid, and Acellular Pertussis (aP) components (DTaP and Tdap) protect against disease. It remains unclear how a parenteral vaccine, which primarily elicits systemic IgG, contributes to protection against a respiratory pathogen. The goal of this study was to investigate mechanisms by which vaccine-induced IgG reaches the respiratory mucosa and contributes to protection against B. pertussis infection. We hypothesized that pertussis-specific systemic IgG is transported from circulation into the airways via the neonatal Fc receptor (FcRn). To test this hypothesis, wild type mice and mice lacking FcRn (FcRn-/-) were immunized with DTaP or passively transferred DTaP-immune serum and challenged with B. pertussis. Post-challenge readouts included kinetics of Pertussis Toxin (PT) IgG in serum and bronchoalveolar lavage fluid (BALf), bacterial load quantification, and histopathology of lung tissues. WT vaccinated mice were able to clear the infection, whereas FcRn-/- vaccinated mice had residual bacterial counts and increased lung inflammation. Passive administration of DTaP-immune sera reduced lung colonization in both WT and FcRn-/- mice. However, FcRn-/- recipients exhibited moderate bronchopneumonia (absent in WT mice). The lower bacterial clearance and exacerbated tissue damage observed in actively and passively immunized FcRn-/- mice was not due to the absence of PT-IgG (or differences in IgG isotype) in BALf. Rather, WT and FcRn-/- mice had similar PT-IgG levels in serum and BALf, suggesting that FcRn-independent mechanisms mediate IgG transport across the lung. PT-IgG progressively increased in BALf of passively immunized FcRn-/- mice post-challenge (along with lung inflammation,) suggesting IgG also diffuses through damaged lung epithelium. We observed that neutrophils from FcRn-/- mice had lower B. pertussis opsonophagocytic capacity as compared to WT. This impairment in IgG-mediated antimicrobial function in the absence of FcRn could explain the increased inflammation in FcRn-/- mice. In conclusion, we have shown that pertussis-specific IgG translocation into the airways appears to be FcRn-independent, and that IgG-mediated B. pertussis neutrophil phagocytosis may contribute to bacterial clearance and tissue preservation post-infection through FcRn interactions.
    • Deciphering relatedness and population demographics in diverse population structures by leveraging haplotype and rare variant sharing detected from whole genome sequencing

      Shetty, Amol Carl; O'Connor, Timothy D; 0000-0001-8790-7649 (2019)
      Genealogical analysis using genomic variants is essential for a variety of applications in human genetics such as estimating population structure, migration events and evolutionary history. The 1000 Genomes Project is an example of a study of human genomic diversity and continental population structure. Multiple studies illustrate the utility of genetic variation for the reconstruction of human migratory patterns within and between continental populations and the demographic events influencing evolutionary history. Current methods for assessment of population structure and genetic relatedness use individual genetic loci and do not take full advantage of the large number of markers provided by whole genome sequencing techniques. More recently, haplotype sharing or identity by descent (IBD) estimates have been used as a promising method to elucidate demographic admixture/migratory events. This dissertation focuses on the application of IBD sharing to decipher genetic relatedness and demographic events that influence population substructure. Knowledge of genome-wide patterns of IBD sharing among individuals helped distinguish between ancient and recent demographic events and detect fine-structure among the recently expanded and admixed New World populations from Peru. This addresses the gap in knowledge regarding the population fine-structure of indigenous and admixed communities from geographically distinct regions of Peru. IBD sharing, primarily utilized to study human demography, was applied to study fine-structure and demography of haploid malarial parasite populations in Southeast Asia which helped elucidate the migratory patterns of the parasite and guide the elimination strategies of the World Health Organization (WHO). Current IBD methods accurately detect long segments based on information from common variants. However, cohorts involving cryptic relatedness mostly share short IBD segments. In light of this limitation, rare variants arising from recent dramatic events of population expansion convey more information on short IBD segments than common variants. This knowledge of IBD sharing leveraged by rare variants influences the timescales at which familial relatedness and population structure can be assessed. In sum, this dissertation illustrates the utility of IBD segments of variable lengths and the accumulation of rare variants within these segments to detect fine-scale population structure at different evolutionary timescales and fills the gaps in knowledge in both human and non-human populations.
    • Elucidation of the Role of CD44 Receptor and its Intracellular Domain in the Progression of Prostate Cancer

      Senbanjo, Linda Temilade; Chellaiah, Meenakshi A. (2019)
      Prostate cancer (PCa) is the second leading cause of cancer-related death in men in the United States partially due to metastatic spread to secondary sites in the bone, brain, lymph nodes and visceral organs. Here, we examine the mechanisms that can facilitate PCa progression/metastasis. CD44 is a multifunctional receptor that functions in tumorigenesis and cancer progression. Recent studies point to CD44 cleavage product; CD44 Intracellular Domain (ICD) as the fragment responsible for transcription of metastasis-related genes (MRGs). Using knockdown and overexpression strategy, we 1) determine the effect of androgen receptor (AR) and CD44 expression on stemness characteristics of PCa cells (PC3), 2) characterize CD44-ICD interaction with RUNX2 in PC3 cells and 3) determine the specificity of interaction of CD44-ICD sequences with RUNX2 and its contribution to PCa progression. We show SOX2 expression is high at mRNA and protein levels of bone metastatic PC3 cells that are AR-negative. Additionally, CD44 regulates the expression of SOX2 and knockdown of SOX2, or CD44 resulted in decrease of cell migration which was reversed with the re-expression of AR. To examine the contribution of CD44-ICD to the tumorigenic potential of PCa cells, we characterized CD44-ICD expression in PCa cells then determined its interaction with RUNX2, a transcription factor also known to promote tumorigenesis. We show the interaction of CD44-ICD/RUNX2 in the nucleus and overexpression of RUNX2 resulted in increased cell migration and tumorsphere formation via the upregulation of MRGs. Consistently, generation and overexpression of CD44-ICD-FL and CD44-ICD carboxyl-terminal deletion constructs resulted in increased interaction with RUNX2 in the nucleus with greater specificity for interacting with constructs: FL and D1 to D3. Likewise, these constructs show an increase in the expression of OPN and MMP-9 genes. However, CD44-ICD-FL, as well as D1 to D3 constructs, shows more specificity to the promoter of MMP-9 in CHIP assay analyses. This also corresponds with the expression levels of MMP-9 protein. These findings not only suggest the specificity of CD44-ICD function towards the expression of MMP-9 but also metastasis relevant events including migration and tumorigenesis. These also implicate CD44-ICD as a novel therapeutic target in cancer cells that express CD44.