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Targeting Heat Shock 27 Kda Protein Induces Androgen Receptor Degradation, Yaxin Li

ETD Archive

Glioblastoma (GBM) is the most common and aggressive brain tumor, with very poor prognosis. Androgen receptor (AR) plays a significant role in the progression of GBM, and anti-androgen agents have the potential to be used for the treatment of GBM. However, AR mutation commonly happens in GBM, which makes the anti-androgen agents less effective. Heat shock 27 kDa protein (HSP27) is a well-documented chaperone protein to stabilize AR. Inhibition of HSP27 results in AR degradation regardless the mutation status of AR, which makes HSP27 a good target to abolish AR in GBM. Identified compound I ((N-(3-((2,5-dimethoxybenzyl)oxy)-4-(methylsulfonamido) phenyl)-4-methoxybenzamide) inhibits GBM cell …

Mechanisms Of Telomere Maintenance In Trypanosoma Brucei, M A G G. Rabbani

ETD Archive

Telomeres are a nucleoprotein structure at the end of the chromosome and are essential for genome integrity and chromosome stability. Telomere lengths are primarily maintained by a telomerase-mediated pathway but can be maintained by a homologous recombination-mediated pathway. However, detailed mechanisms of telomere maintenance are still unclear in many eukaryotes, including an important human pathogen, Trypanosoma brucei. Telomeres can be elongated by telomerase in T. brucei, a causative agent of fatal sleeping sickness in humans and nagana in cattle. T. brucei evades host immune response by regularly switching its major surface antigen, variant surface glycoprotein (VSG), a process known as …

Biochemical Characterization Of Induced Pluripotent Stem Cell-Derived Cardiomyocytes As A Model Of Barth Syndrome, Alisha J. House

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Barth Syndrome (BTHS) is an X-linked inborn error of metabolism (IEM) which manifests as a multi-systemic disease. One of the primary symptoms is dilated cardiomyopathy, and alongside the cardiovascular disease that arises, patients often experience metabolic abnormalities such as 3-methylglutaconic aciduria, growth retardation, and neutropenia. There has been a need for the development of a suitable in vitro modeling system which will accurately recapitulate the biochemical and physical nature of BTHS. The purpose of this project has been to develop a model for studying the biochemical pathogenesis of Barth Syndrome using human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs). To achieve …

Evaluation Of A Small Molecule Agonist Of Epha2 Receptor Tyrosine Kinase And Copalic Acid Analogs As Prostate Cancer Therapeutics, Nethrie Idippily

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Project I: Chemotherapeutic drugs have many side effects that are undesirable and are highly toxic. Therefore, there is a growing need for the development of drugs with enhanced efficacy, specificity, and potency to provide cancer patients with a better prognosis. It was discovered that a member of the Receptor Tyrosine Kinase family, EphA2, may prove to be a viable target in developing anti-cancer agents. In the presence of its ligand, EphA2 receptor is responsible for apoptotic and anti-migratory activity. However, in the absence of ligand, EphA2 is able to stimulate cell migration and therefore tumorigenic activity. These conflicting roles of …

Quantitative Analysis Of Bleomycin In Rat Plasma By Lc-Ms/Ms, Huawen Li

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Bleomycin is the most commonly used compound in its group of antineoplastic drugs. It works on tumor cells by single and double stranded DNA cleavage after its activation, in which it blocks tumor cells’ DNA replication or transcription activities to inhibit tumor cells’ growth. Bleomycin sulfate (Blenoxane) is the most popular preparation used in clinical research, and contains Bleomycin fractions of A2 and B2, which causes difficulties in quantitative analysis. This work uses the metal chelating property of Bleomycin as an advantage to simplify and improve sensitivity of existing quantitative methods. Copper was spiked in excess to plasma samples, followed …

Computational Investigation Of Protein Assemblies, Sm Bargeen Alam Turzo

ETD Archive

Selective nitrosylation of glyceraldehyde 3-phosphate dehydrogenase (GAPDH) at Cys-247 affects gene regulation through the interferon-gamma (IFN- γ) activated inhibitor of translation (GAIT) complex. Oxidized low-density lipoprotein (LDL_ox) and INF-γ induce assembly of the nitrosylase complex composed of inducible nitric oxide synthase (iNOS), S100A8, and S100A9 proteins. Crystal structure of the complex of GAPDH and S100A8A9 is not known, structural prediction method were employed by protein-protein docking and binding energy calculation with PatchDock and FIREDock respectively. Candidate models were selected, based on a weight factor calculated, from the computational method developed from the "artificial protease" cleavage mapping Fe(III) (s)-1-(p- …

Il-17 Drives Copper Uptake And Activation Of Growth Pathways In Colorectal Cancer Cells In A Steap4-Dependent Manner, Evan Martin

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Colorectal cancer is a disease characterized by abnormal, invasive cell growth beginning in the colon or rectum. The third most common type of cancer worldwide, approximately one million new cases of the disease are diagnosed across the globe annually, resulting in an estimated 700,000+ deaths. One major risk factor associated with development of colorectal cancer is the presence of chronic inflammation in the large intestine, also known as colitis. Inflammation is a complex immune response against harmful stimuli, characterized by symptoms including heat, redness, swelling and pain. One important molecular mediator of this process is interleukin 17 (IL-17), a pro-inflammatory …

Il-17 Drives Copper Uptake And Activation Of Growth Pathways In Colorectal Cancer Cells In A Steap4-Dependent Manner, Evan Martin

ETD Archive

Colorectal cancer is a disease characterized by abnormal, invasive cell growth beginning in the colon or rectum. The third most common type of cancer worldwide, approximately one million new cases of the disease are diagnosed across the globe annually, resulting in an estimated 700,000+ deaths. One major risk factor associated with development of colorectal cancer is the presence of chronic inflammation in the large intestine, also known as colitis. Inflammation is a complex immune response against harmful stimuli, characterized by symptoms including heat, redness, swelling and pain. One important molecular mediator of this process is interleukin 17 (IL-17), a pro-inflammatory …

Therapeutic Potential Of Rhtrail For Malignant Melanoma, Katherine Ann Turner

ETD Archive

The application of recombinant human Tumor Necrosis Factor-Related Apoptosis-Inducing Ligand (rhTRAIL) for the treatment of cancer holds great promise due to its ability to selectively induce apoptosis in cancer cells while not harming normal healthy cells. This is evident by the robust levels of apoptosis induced in malignant melanoma cells while no event of apoptosis was observed in the non-transformed counterpart of melanomas, melanocytes. However, the clinical utility of rhTRAIL is limited due to the heterogeneity seen in rhTRAIL-sensitivity among cancers. rhTRAIL-resistance is especially prevalent in cases of malignant melanoma. Melanoma rhTRAIL-resistance can be attributed to a number of different …

Using Nucleotide Analogs As Biochemical Probes To Evaluate The Mechanisms Involved In Translesion Replication By A High Fidelity Dna Polymerase, Anvesh Dasari

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Translesion DNA synthesis (TLS) allows DNA polymerases to incorporate nucleotides opposite and beyond damaged DNA. This activity is an important risk factor for the initiation and progression of genetic diseases including cancer. My study evaluates the ability of a high-fidelity DNA polymerase to perform TLS with 8-oxo-guanine, a pro-mutagenic DNA lesion formed by reactive oxygen species. Using modified purine and non-natural indole analogs as biochemical probes, I have evaluated the influence of desolvation, hydrogen bonding interactions, and shape complementarity towards nucleotide binding and incorporation opposite the miscoding lesion 8-oxo-guanine by the high fidelity gp43exo- DNA polymerase. In Chapter II, I …

The Regulatory Role Of Interleukin-1 Receptor Associated Kinase M In Toll-Like Receptor Signaling, Hao Zhou

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Toll-like receptors (TLRs) are one of the major groups of pattern recognition receptors. TLRs are able to recognize the pathogen-associated molecular patterns and transduce their signals through the adaptor molecule MyD88 and members of the IL-1R-associated kinase family (IRAK1, 2, M and 4). IRAKM was previously known to function as a negative regulator that prevents the dissociation of IRAKs from MyD88, thereby inhibiting downstream signaling. However, we now found that IRAKM was also able to interact with MyD88-IRAK4 to form IRAKM Myddosome to mediate TLR7-induced MEKK3-dependent second wave NF¿B activation. As a result, the IRAKM-dependent pathway only induced expression of …

Profiling Cell Surface Sialylation And Desialylation Dynamics Of Immune Cells, Dan Wang

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Sialic acids (SAs) are a diverse family of naturally occurring 2-keto-3-deoxy-nononic acids that are involved in a wide range of biological processes, including early fetal development, cellular recognition, and utilization by microbes. While it is clear that cell surface SAs are highly involved in the immune system, the sialylation status of individual immune cells and functions are still unknown. In this study, I combined the newly developed LC-MS/MS methods with flow cytometry and confocal microscopy to systematically study the sialylation and desialylation dynamics of macrophages at different conditions.

First, I developed an accurate LC-MS/MS method to quantify free SA in …

Study Of Electron Transfer Through The Reductase Domain Of Neuronal Nitric Oxide Synthase And Development Of Bacterial Nitric Oxide Synthase Inhibitors, Yue Dai

ETD Archive

Crystal structure of neuronal Nitric Oxide Synthase reductase (nNOSr) implies that large-scale domain motion is essential for electron transfer. However, the details are not well understood. To address this, we generated a functioning “Cys-lite” version of nNOSr and then replaced the nNOSr Glu816 and Arg1229 residues with Cys in the FMN and FAD domains (CL5SS) in order to allow cross-domain disulfide bond formation under pH 9 or to cross-linking using bis-maleimides. Cross-linked CL5SS exhibited a =95% decrease in cytochrome c reductase activity and reduction of the disulfide bond restored the activities. The results demonstrate that a conformational equilibrium involving FMN …

Time And Temperature Dependent Surface Tension Measurements Of Responsive Protein-Based Polymer Surfactant Solutions, Hakan Celik

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A three-armed star elastin-like polypeptide (ELP-foldon) has thermoreversible character which exhibits phase separation above a transition temperature (Tt) in physiologic salt concentrations. At lower salt concentration, the ELP-foldon behaves like a thermoresponsive surfactant, exhibiting micelle formation above its Tt. The purpose of this study is characterize the surfactant behavior of the ELP-foldon at air-liquid interface by measuring the surface tension. The surface tension is measured as a function of time for different ELP concentrations from 10 nM to 50 ℗æM and over range of temperatures from 25 to 35 ℗¿ using the axisymmetric drop shape analysis (ADSA). The ADSA is …

The Role Of Platelets In Hyaluronan Degradation, Sami Albeiroti

ETD Archive

Following tissue injury, platelets rapidly interact with the exposed extracellular matrix (ECM) of blood vessel wall and the surrounding tissues. Hyaluronan (HA) is a major polysaccharide component of the ECM and plays a significant role in regulating inflammation. Human platelets can degrade HA from the surfaces of activated endothelial cells into fragments capable of inducing immune responses by monocytes. In addition, platelets contain the enzyme hyaluronidse-2 (HYAL2), one of two major hyaluronidases that digest HA in somatic tissues. The deposition of HA increases in the inflamed tissues in several inflammatory diseases, including Inflammatory Bowel Disease (IBD). This study provides evidence …

Role Of Mtorc1 And Autophagy In The Regulation Of Chemoresistance In Leukemic B-Cells, Arishya Sharma

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Deregulated mTORC1 contributes to tumorigenesis and chemoresistance. However, mTORC1-specific inhibitors (rapalogs) show modest efficacy in the clinic, as they unleash the feedback inhibition on upstream, prosurvival pathways. An alternative approach is to target downstream functions of mTORC1. We investigated acquired resistance to fludarabine (Flu), a purine analog, active agent for chronic lymphocytic leukemia (CLL). Elevated phospho-p70S6k (T389), an mTORC1 activation marker, predicted Flu resistance in a panel of B-cell tumor lines, and primary CLL cells. However, the rapalogs induced moderate cell death in Flu-resistant (FluR) and primary CLL cells. Activation of oncogenic pathways, including mTORC1, induces profound metabolic reprogramming to …

Combinatorial Activation Of Stat3 By Egf And Thrombin In Endothelial Cells, Matthew S. Waitkus

ETD Archive

Endothelial cells line the luminal surface of blood vessels and form a regulatory interface between the bloodstream and underlying tissues. The endothelium responds to diverse, and potentially conflicting, environmental signals to regulate vessel growth, leukocyte adhesion, thrombogenicity, and vascular tone. Signaling pathways may interact, or "crosstalk," in combinatorial signaling environments to enable cells to process disparate extracellular information at downstream signaling nodes and formulate appropriate biological responses based on combinations of extracellular stimuli. We have reported that simultaneous stimulation of endothelial cells with EGF and thrombin synergistically induces expression of immediate early genes (IEGs) associated with growth and angiogenesis. We …

The Role Of Rnase L In Thymic Homeostasis And Humoral Responses, Lin Zhang

ETD Archive

RNase L is highly expressed in the spleen, thymus, and multiple immune cells, suggesting that it may play an important role in the immune system against microbes. Previous studies in the lab have shown that deficiency of RNase L results in enlarged thymuses in mice at young age. However, relatively little is known about its influence on the thymic development and adaptive immunogenicity. The present Ph.D. study focuses on investigating the role of RNase L in the thymic homeostasis and humoral immune responses, thereby gaining new insights into the molecular aspects in thymocyte development, maturation and adaptive immunity.By using RNase …

Conformational Regulation Of Membrane Localization And Activation Of Talin, Pallavi Dwivedi

ETD Archive

Talin is a cytosolic protein which is known to be one of the key players involved in integrin mediated cell adhesion dependent processes, including blood coagulation, and tissue remodeling. It connects the extracellular matrix with the actin cytoskeleton. Talin comprises of a head domain (talin-H) and a rod domain (talin-R). Talin-H is further subdivided in F0, F1, F2 and F3 domains. Talin-R contains 13 contiguous helical bundle domains (R1-R13) followed by an actin binding dimerization domain (DD). The F3 domain contains a key integrin binding site that regulates integrin activation. In our previous studies, we have shown that cytosolic talin …

Design, Synthesis And Characterization Of Chain-End Functionalized Glyco-Polymer For Efficient Proteomic Analysis, Satya Nandana Narla

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No abstract provided.

Rnase L Manipulates Macrophages In Innate Immunity And Tumor Growth, Xin Yi

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RNase L is one of the key enzymes in the 2-5A system of interferon (IFN) action against viral infection and cellular proliferation. Tissue distribution analysis has revealed that RNase L is highly expressed in the spleen, thymus, lung, testis, intestine and most of immune cells such as T, B cells and macrophages. However, the physiological role of RNase L in the immune system is largely unknown. My thesis thus focused on studying the possible physiological role of RNase L in macrophages. By using bone marrow-derived macrophages (BMMs) from RNase L+/+and -/- mice, we demonstrated that RNase L is involved in …

Structural Basis Of Lman1 Cargo Capture In Er & Release In Ergic, Vaijayanti Das

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No abstract provided.

Molecular Basis Of The Role Of Kindlin 2 In Cell Adhesion, Hettiarachchige Dhanuja Deepamalee Perera

ETD Archive

Kindlins are a novel family of cytoskeleton proteins that plays an important role in the activation of heterodimeric integrin transmembrane receptors. These receptors play a vital role in cell adhesion, cell migration and other cellular processes. The Kindlin protein family consists of three homologous proteins, Kindlin 1, 2 and 3 with differential tissue distribution and varying expression levels in human. Mutations of Kindlin 1 protein are clinically linked to Kindler Syndrome. Kindlin 2 knockdown studies have shown embryonic lethality in mouse, but no diseases have been clinically testified. Kindlin 3 mutations are associated with bleeding disorders, e.g. LADIII. Talin is …

Structural Studies Of Talin-Mediated Integrin Activation, Esen Bakhautdin

ETD Archive

Activation of heterodimeric (alpha/beta) integrin transmembrane receptors by the cytoskeletal protein talin is essential for many important cell adhesive responses including cell-extracellular matrix contact, cell motility and survival. A key step in this process involves interaction of phosphotyrosine-binding (PTB) domain in the N-terminal head of talin (talin-H) with integrin beta3membrane-proximal cytoplasmic tails (beta-MP-CTs). Compared to talin-H, intact talin exhibits low potency in inducing integrin activation. Using TROSY NMR spectroscopy, we found that the large C-terminal rod domain of talin (talin-R) interacts with talin-H and allosterically restrains talin in a closed conformation. Structural, mutational, and biochemical analyses demonstrate that talin-R specifically …