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Regulation Of Egf Receptor Dynamics By Protein Tyrosine Phosphatases, Calixte Monast Jan 2013

Regulation Of Egf Receptor Dynamics By Protein Tyrosine Phosphatases, Calixte Monast

Publicly Accessible Penn Dissertations

The phosphorylated epidermal growth factor receptor (EGFR) initiates intracellular signaling processes that regulate cell growth, survival, and migration, and disregulated EGFR-mediated signaling occurs in many cancers. While the processes that lead to EGFR activation and phosphorylation have been studied in detail, quantitative aspects of the spatiotemporal regulation of EGFR by protein tyrosines phosphatases (PTPs) are not well understood. To begin to address this, we developed a new compartmentalized mechanistic model of EGFR phosphorylation dynamics and used it to interpret quantitative biochemical measurements to show that EGFR is dephosphorylated at the plasma membrane and in the cell interior with a time ...


Mechanical Development And Functional Mechanosensitivity During Early Cardiogenesis, Stephanie Feldman Majkut Jan 2013

Mechanical Development And Functional Mechanosensitivity During Early Cardiogenesis, Stephanie Feldman Majkut

Publicly Accessible Penn Dissertations

This thesis addresses the questions of when and how mechanical stiffness arises during embryonic heart development and how mechanics affects early cardiomyocyte and myocardium contractile function and cytoskeletal organization. Previous studies addressing how mechanics influence the contractile and electrochemical capacity of mature cardiomyocytes on compliant substrates are reviewed in light of theory explaining how contractile striated fibers might optimally align on intermediate substrates. Embryonic heart and brain tissue stiffness through early development are measured by micropipette aspiration, and the earliest functional heart is found to be three-fold stiffer than early embryonic tissue while brain remains soft. Contraction strain in intact ...


Control Of The Tumor Suppressor P53 By Regulating Mdm2 Activity And Stability, Ruchira S. Ranaweera Jan 2013

Control Of The Tumor Suppressor P53 By Regulating Mdm2 Activity And Stability, Ruchira S. Ranaweera

Publicly Accessible Penn Dissertations

p53 is a tumor suppressor that is widely mutated or deleted in cancer cells. Mdm2, an E3 ubiquitin ligase, is the master regulator of p53. It targets p53 for proteasomal degradation, restraining the potent activity of p53 and enabling cell survival and proliferation. There are complex regulatory mechanisms balancing the activity and stability of Mdm2 in a cell. Mdm2 has an extremely short half-life in the unstressed cell and its regulation is not well understood. Like most E3 ligases, Mdm2 can autoubiquitinate. Previously, the sole function of autoubiquitination was thought to be to signal Mdm2 degradation. Here I show that ...


Abnormal Smooth Muscle Contraction Alters Gut Motility And Propagates Epithelial Invasion In The Larval Zebrafish Intestine, Joshua Abrams Jan 2013

Abnormal Smooth Muscle Contraction Alters Gut Motility And Propagates Epithelial Invasion In The Larval Zebrafish Intestine, Joshua Abrams

Publicly Accessible Penn Dissertations

Coordinated smooth muscle contraction is critical for force production and proper functioning of numerous organ systems. Activation at the myosin motor domain via phosphorylated myosin light chain (phospho-MLC) remains the primary signal to initiate contraction, but it is now appreciated that there are additional force modulators also present in smooth muscle. One particularly well studied modulatory protein is Caldesmon (CaD), which has been implicated in controlling contractile force in vascular smooth muscle, however little is known of CaD's physiological role in vivo. Studies in vitro have shown that CaD inhibits actomyosin interactions and that this effect is reversed after ...


Jip1 Regulates Axonal Transport Of App And Autophagosomes Via Coordination Of Kinesin And Dynein Motors, Meng-Meng Fu Jan 2013

Jip1 Regulates Axonal Transport Of App And Autophagosomes Via Coordination Of Kinesin And Dynein Motors, Meng-Meng Fu

Publicly Accessible Penn Dissertations

Neurons are specialized cells that extend polarized processes called dendrites and axons in order to maintain synaptic connections over long distances. Consequently, neuronal homeostasis requires axonal transport of organelles, such as mitochondria, synaptic vesicles, and autophagosomes. The microtubule-based motors responsible for long-distance fast axonal transport are the anterograde kinesin motors and the retrograde dynein motors. Two cargos that exhibit robust axonal transport characterized by high speeds with few directional switches are APP- (amyloid precursor protein) positive vesicles and autophagosomes. While APP-positive vesicles transport occurs in both anterograde and retrograde directions, autophagosomes move unidirectionally in the retrograde direction. Here, we demonstrate ...


The Interplay Between Lewy Body-Like Alpha-Synuclein Aggregates Nd Protein Degradation Pathways In Cell-Based Model Of Parkinson's Disease, Selcuk Aski Tanik Jan 2013

The Interplay Between Lewy Body-Like Alpha-Synuclein Aggregates Nd Protein Degradation Pathways In Cell-Based Model Of Parkinson's Disease, Selcuk Aski Tanik

Publicly Accessible Penn Dissertations

Cytoplasmic alpha-synuclein (a-syn) aggregates, including Lewy bodies (LBs), are pathological hallmarks of a number of neurodegenerative diseases, most notably Parkinson's disease (PD). Activation of intracellular protein degradation pathways (Pdps) to eliminate these aggregates has been proposed as a therapeutic approach for PD and other synucleinopathies, but the interplay between LB-like a-syn aggregates and Pdps is not completely understood. Here, we investigate this interplay by utilizing a recently developed cellular model in which intracellular LB-like a-syn inclusions accumulate after delivery of pre-formed a-syn fibrils (Pffs) into a-syn-expressing HEK293 cells or cultured primary neurons. This thesis describes the interplay between LB-like ...


The Role Of Tipe2 In The Regulation Of Inflammation And Tumorigenesis, Derek Johnson Jan 2013

The Role Of Tipe2 In The Regulation Of Inflammation And Tumorigenesis, Derek Johnson

Publicly Accessible Penn Dissertations

TIPE2 is a recently discovered regulator of immunity and inflammation. Here we describe a new function of TIPE2 in the regulation of Ras signaling and Tumorigenesis. By using various stimuli and inhibitors in T Cells and macrophages we discovered that TIPE2 is regulated at both the message and protein level by inflammatory stimuli. TIPE2 mRNA is regulated in the short to intermediate term by an NF-Kappa B induced micro RNA, and TIPE2 is also ubiquitylated and degraded, possibly by SCF-Beta TRCP. Mechanistically TIPE2 interacts with and inhibits the Ras-interacting domain of the RalGDS family of Ras effectors, leading to a ...


Epigenetic Basis For The Oncogenic Potential Of Idh Mutations, Chao Lu Jan 2013

Epigenetic Basis For The Oncogenic Potential Of Idh Mutations, Chao Lu

Publicly Accessible Penn Dissertations

Although many complex diseases including cancer manifest aberrant cellular metabolism and chromatin structure, the molecular connection between two processes remains poorly understood. The metabolite, α-ketoglurate (αKG), is a critical co-factor for a number of chromatin modifying enzymes. Its structural analogue, 2-hydroxyglutarate (2HG), was recently identified as the product of cancer-associated mutations in isocitrate dehydrogenases (IDH). To determine whether metabolic perturbation can disrupt chromatin remodeling and transcription, I investigated the epigenetic consequences of 2HG-producing IDH mutations. In this thesis, 2HG was demonstrated to be a competitive inhibitor for αKG-dependent chromatin modifiers including TET family DNA hydroxylases and jumonji-C histone demethylases. Expression ...


Host-Apicomplexan Parasite Interactions: Leveraging Biological Discovery Into Antiparasitic Drug Development, Melanie Grace Millholland Jan 2013

Host-Apicomplexan Parasite Interactions: Leveraging Biological Discovery Into Antiparasitic Drug Development, Melanie Grace Millholland

Publicly Accessible Penn Dissertations

The obligate intracellular pathogens Plasmodium falciparum and Toxoplasma gondii remodel their host cell to facilitate their intracellular development and progress through their asexual life cycle, a virulent lytic cycle responsible for parasite-mediated pathogenesis. While several studies have highlighted parasite proteins that interact with the host cell during this cycle, host proteins exploited by the parasite for successful growth and conversely, host molecules evolutionarily tuned to control parasite infection remain unclear. We addressed this question from both sides of the host-parasite interaction in the hope to leverage biological discovery of host molecules involved in infection into the validation of novel drug ...


Discovering Novel Intrinsic Antiviral Responses To Arboviruses: From Transcription To Intestinal Innate Immunity, Jie Xu Jan 2013

Discovering Novel Intrinsic Antiviral Responses To Arboviruses: From Transcription To Intestinal Innate Immunity, Jie Xu

Publicly Accessible Penn Dissertations

Many (re) emerging pathogens are arthropod-borne, transmitted via an insect vector, and cause significant health and agricultural problems worldwide. Despite their significance, there are few vaccines and no targeted therapies that exist. This is at least in part due to our limited understanding of virus-host interactions and the mechanisms used by hosts to restrict infection. In particular, insect vectors play a critical role in the transmission and spread of these pathogens, but performing molecular and genetic studies has proven to be difficult. Drosophila is a model organism that shares a high degree of conservation with insect vectors and has a ...


Mitotic Kinases In Meiosis, Olga Davydenko Jan 2013

Mitotic Kinases In Meiosis, Olga Davydenko

Publicly Accessible Penn Dissertations

The goal of the first meiotic division is the faithful segregation of homologous chromosomes to produce a gamete with proper ploidy. To achieve this, chromosomes must align at the metaphase plate of a bipolar meiotic spindle, biorient and separate equally at anaphase. Compared to mitosis, meiotic divisions are associated with extra challenges for the mitotic cell division machinery. First, meiotic spindles are acentrosomal, and bipolarity establishment is more complex than during mitosis. Second, homologous chromosomes must biorient and separate during meiosis I, unlike during most other cell divisions, which separate sister chromatids. Meiotic cells have adapted the mitotic enzymatic machinery ...


Molecular Mechanisms Of Alternative Splicing Regulation: An Investigation Of The Spliceosome Repressed By Hnrnp L On Cd45 Exon 4, Ni-Ting Chiou Jan 2013

Molecular Mechanisms Of Alternative Splicing Regulation: An Investigation Of The Spliceosome Repressed By Hnrnp L On Cd45 Exon 4, Ni-Ting Chiou

Publicly Accessible Penn Dissertations

Alternative splicing is a key step in gene regulation and involves the differential selection of splice sites to generate different pre-mRNA transcripts. It has been shown that 90-95% of pre-mRNAs are alternatively spliced in human cells. Pre-mRNA splicing is catalyzed the spliceosome, which consists mainly of the U1, U2, U4, U5 and U6 snRNP, and about a hundred of non-snRNP proteins. Splicing regulators that bind to enhancer or silencer elements on the pre-mRNA can alter assembly of these spliceosome components. Understanding how splicing regulators control spliceosome assembly will bring insights to the prediction of splice site choices. In our lab ...


Bridging The Gap: Defining The Molecular Mechanisms Of Cep290 Disease Pathogenesis, Theodore George Drivas Jan 2013

Bridging The Gap: Defining The Molecular Mechanisms Of Cep290 Disease Pathogenesis, Theodore George Drivas

Publicly Accessible Penn Dissertations

Mutations in the gene CEP290 cause an array of debilitating and phenotypically distinct human diseases, ranging in severity from the devastating blinding disease Leber congenital amaurosis (LCA) to Senior Løken Syndrome, Joubert syndrome, and the embryonically lethal Meckel-Grüber syndrome. The pathology observed in these diseases is thought to be due to CEP290's essential role in the development and maintenance of the primary cilium, but despite its critical role in biology and disease we know only little about CEP290's function. Here we identify four novel functional domains of the protein, showing that CEP290 directly binds to cellular membranes ...


Determining The Role Of Autophagy In Her2/Neu-Induced Mammary Tumor Dormancy And Recurrence, Samantha L. Dwyer Jan 2013

Determining The Role Of Autophagy In Her2/Neu-Induced Mammary Tumor Dormancy And Recurrence, Samantha L. Dwyer

Publicly Accessible Penn Dissertations

Mortality from breast cancer is principally due to tumor recurrence. Recurrent breast cancers arise from the pool of residual tumor cells, termed minimal residual disease, that survive treatment and may exist in a dormant state for up to 20 years following treatment of the primary tumor. As recurrent breast cancer is typically a fatal disease, understanding the mechanisms underlying dormant tumor cell survival is a critical priority in breast cancer research. Using a HER2/neu inducible bitransgenic mouse model, we demonstrate that the survival and recurrence of dormant mammary tumor cells is dependent on autophagy. We find that autophagy is ...


Regulation Of T Cell Receptor Signaling By Diacylglycerol Kinases And Phosphatidylinositol Transfer Proteins, Rohan Prakash Joshi Jan 2013

Regulation Of T Cell Receptor Signaling By Diacylglycerol Kinases And Phosphatidylinositol Transfer Proteins, Rohan Prakash Joshi

Publicly Accessible Penn Dissertations

Signals transduced through the T cell receptor (TCR) lead to T cell differentiation, proliferation, and elaboration of cytokines, all of which are required for optimal immunity. Phosphoinositide (PI) mediated signaling plays a particularly prominent role in this process. TCR signaling is amplified by the activation of phospholipase C γ1 (PLCγ1), which cleaves phosphatidylinositol-4,5-bisphosphate (PIP2) to form the second messengers diacylglycerol (DAG) and inositol triphosphate (IP3). Regulation of PI and products such as DAG are therefore essential for normal TCR signaling. DAG levels are reduced by diacylglycerol kinases (DGKs), which metabolize DAG and diminish DAG-mediated signaling. In T ...


Cd8+ T-Cell Responses In A Th2-Polarized Inflammatory Setting, Vanessa Christina Kurzweil Jan 2013

Cd8+ T-Cell Responses In A Th2-Polarized Inflammatory Setting, Vanessa Christina Kurzweil

Publicly Accessible Penn Dissertations

A variety of external factors can affect the activation status, effector function, and memory differentiation of T cells. Such factors include resident microbiota and the cytokine milieu produced in a host. Loss of tolerance to commensal bacteria and excess inflammatory cytokines such as IL-4 have been implicated in the pathogenesis of many diseases, including inflammatory bowel disease (IBD) and allergy. Although these diseases are traditionally associated with CD4+ T cells, it is increasingly appreciated that CD8+ T cells in these settings may either contribute to or reduce pathology. This work explores the influence of various factors on CD8+ T-cell responses ...


Genome-Wide Analysis Of Rna Secondary Structure In Eukaryotes, Fan Li Jan 2013

Genome-Wide Analysis Of Rna Secondary Structure In Eukaryotes, Fan Li

Publicly Accessible Penn Dissertations

The secondary structure of an RNA molecule plays an integral role in its maturation, regulation, and function. Over the past decades, myriad studies have revealed specific examples of structural elements that direct the expression and function of both protein-coding messenger RNAs (mRNAs) and non-coding RNAs (ncRNAs). In this work, we develop and apply a novel high-throughput, sequencing-based, structure mapping approach to study RNA secondary structure in three eukaryotic organisms.

First, we assess global patterns of secondary structure across protein-coding transcripts and identify a conserved mark of strongly reduced base pairing at transcription start and stop sites, which we hypothesize helps ...


Regulators Of Mouse And Human Beta Cell Proliferation, Yang Jiao Jan 2013

Regulators Of Mouse And Human Beta Cell Proliferation, Yang Jiao

Publicly Accessible Penn Dissertations

Diabetes mellitus is an increasingly prevalent metabolic disorder that is estimated to affect over 300 million people by 2025. Common to either type 1 or type 2 diabetes is a progressive inadequacy of functional beta-cell mass. Recent studies have shown that during times of prolonged metabolic demand for insulin, the endocrine pancreas can respond by increasing beta-cell mass by beta-cell proliferation. Advances that further our knowledge of the molecular factors that control beta-cell proliferation will be crucial for understanding the homeostasis of beta-cell mass during adulthood, and are pivotal for any attempt to use instructive cues to induce the proliferation ...


Functions Of Atoh8 And Mekk3 In Cardiovascular Development, David Robert Rawnsley Jan 2013

Functions Of Atoh8 And Mekk3 In Cardiovascular Development, David Robert Rawnsley

Publicly Accessible Penn Dissertations

The transcription factor ATOH8 and the kinase MEKK3 are evolutionary conserved proteins with known expression within the cardiovascular system. However, the roles of these proteins in the developing heart are undefined. We used a combination of mutant mouse models, morpholino gene suppression in the zebrafish, and cell culture to determine the role of these two proteins in cardiovascular system. Our experiments with ATOH8 reveal a direct interaction between ATOH8 and the cardiovascular transcription factors FOG2 and GATA4. This interaction is required in vivo in the developing zebrafish heart, where Atoh8 functions with Gata and Fog factors to promote proper cardiac ...


Arx Influences Cortical Function By Regulating Progenitor Cell Proliferation, Jacqueline Christine Simonet Jan 2013

Arx Influences Cortical Function By Regulating Progenitor Cell Proliferation, Jacqueline Christine Simonet

Publicly Accessible Penn Dissertations

Mutations in the Aristaless-related homeobox (ARX) gene are found in a spectrum of epilepsy and X-linked intellectual disability disorders in children. During development Arx is expressed in pallial ventricular zone (VZ) progenitor cells which give rise to the excitatory projection neurons of the cortex. Arx-/Y mice were shown to have decreased proliferation in the cortical VZ resulting in smaller brains; however, the basis for this reduced proliferation was not established. To determine the role of ARX on cell cycle dynamics in cortical progenitor cells, we generated cerebral cortex specific Arx mouse mutants (cKO). The loss of pallial Arx resulted ...


The Role Of Arx In Specification And Maintenance Of Pancreatic Islet Α-Cells, Crystal Ley Wilcox Jan 2013

The Role Of Arx In Specification And Maintenance Of Pancreatic Islet Α-Cells, Crystal Ley Wilcox

Publicly Accessible Penn Dissertations

Correct specification and maintenance of pancreatic islet cells is an intricate process. Previous studies have demonstrated the essential role transcription factors play in this process. For islet glucagon-producing alpha-cells one such transcription factor is the aristaless-related homeobox gene (Arx). Previous studies have demonstrated that Arx is necessary and sufficient for alpha-cell specification where ablation of Arx results in complete loss of the alpha-cell lineage and misexpression leads to conversion into an alpha-cell phenotype. However, the role of Arx in maintenance of alpha-cell fate as well the impact non-null mutations of Arx have on alpha-cell development has not been explored. In ...


Cellular Mechanisms Of Mammalian Liver Regeneration, Kilangsungla Yanger Jan 2013

Cellular Mechanisms Of Mammalian Liver Regeneration, Kilangsungla Yanger

Publicly Accessible Penn Dissertations

ABSTRACT

CELLULAR MECHANISMS OF MAMMALIAN LIVER REGENERATION

Kilangsungla Yanger

Ben Z. Stanger

The liver is an essential organ that aids in metabolic processes, protein synthesis and detoxification of harmful substances. As the centre for detoxification, the liver is able to compensate for this routine damage with its robust regenerative ability. All vertebrate livers, for example, can make up for tissue mass loss (via surgical excision of a portion of the liver) by replication of their differentiated cells within the remnant lobes. These differentiated cells include parenchymal cells such as the hepatocytes and biliary epithelial cells (BECs) and also non-parenchymal cells ...


Local Cytoskeletal And Organelle Interactions Impact Molecular Motor-Driven Early Endosomal Trafficking, Allison Lorraine Zajac Jan 2013

Local Cytoskeletal And Organelle Interactions Impact Molecular Motor-Driven Early Endosomal Trafficking, Allison Lorraine Zajac

Publicly Accessible Penn Dissertations

Molecular motors generate the force needed for long-distance transport of cargos and organelles in the cell. How motor proteins attach to a diverse array of cargos and navigate to the correct location in the cell with enough fidelity to maintain organelle integrity is only starting to be understood. Studying the properties of individual motors, and their fine-tuning by regulatory molecules, is one area of active investigation in vitro. However, the organization of the cell, and the variability of the environment within a single cell, cannot be fully reconstituted in vitro. We investigated the effects of the crowded intracellular environment on ...


Regulation And Dynamic Behavior Of The Heat Shock Transcription Factor Hsf-1 In C. Elegans, Elizabeth A. Morton Jan 2013

Regulation And Dynamic Behavior Of The Heat Shock Transcription Factor Hsf-1 In C. Elegans, Elizabeth A. Morton

Publicly Accessible Penn Dissertations

Eukaryotic cells respond to heat stress by activating the transcription factor HSF1. In addition to its role in stress response, HSF1 also functions in protein homeostasis, aging, innate immunity, and cancer. Despite prominent HSF1 involvement in processes pertinent to human health and disease, there are still gaps in our understanding of HSF1. For example, controversy exists regarding the localization of HSF1, the identity of HSF1 regulators, and the function and conservation of heat-induced HSF1 stress granules. Many of the physiological roles for HSF1 have been defined using the model organism Caenorhabditis elegans, yet little is known about how the molecular ...


Camp Promotes Retinal Midline Crossing At The Zebrafish Optic Chiasm, Alison Dell Jan 2013

Camp Promotes Retinal Midline Crossing At The Zebrafish Optic Chiasm, Alison Dell

Publicly Accessible Penn Dissertations

During development, axons navigate a complex intracellular milieu, often over long distances to reach target tissues at stereotyped locations. The basis for this navigation lies in the growth cone - a motile structure at the tip of the growing axon. Studded with receptors for an array of guidance cues, the growth cone interprets a balance of attractive and repellant signals en route to its target. However, during pathfinding, axons often must extend along or near areas expressing repellents. We previously showed that growth cones can modulate their response to multiple repellents through a G protein coupled receptor, calmodulin, cAMP and PKA ...


Developmental Regulation Of Strongyloides Stercoralis Infectious Third-Stage Larvae By Canonical Dauer Pathways, Jonathan David Chaffee Stoltzfus Jan 2013

Developmental Regulation Of Strongyloides Stercoralis Infectious Third-Stage Larvae By Canonical Dauer Pathways, Jonathan David Chaffee Stoltzfus

Publicly Accessible Penn Dissertations

Parasitic nematodes inflict a vast global disease burden in humans as well as animals and plants of agricultural importance; understanding how these worms infect their hosts has significant health and economic implications. In humans, soil-transmitted parasitic nematodes cause hookworm disease and strongyloidiasis, and vector-transmitted parasitic nematodes cause filariasis. The infectious form of the species causing these diseases is a developmentally arrested third-stage larva (L3i). Molecular mechanisms governing L3i developmental arrest and activation within a host have been poorly understood. An analogous developmentally arrested third-stage larva--the dauer larva--forms during stressful environmental conditions in the free-living nematode Caenorhabditis elegans and is controlled ...


The Multifunctional Protein Daxx: Studies Of Its Biology And Regulation, And Discovery Of A Novel Function, Trisha Agrawal Jan 2013

The Multifunctional Protein Daxx: Studies Of Its Biology And Regulation, And Discovery Of A Novel Function, Trisha Agrawal

Publicly Accessible Penn Dissertations

Daxx, a multifunctional protein with a diverse set of proposed functions, is ubiquitously expressed and highly conserved through evolution. A primarily nuclear protein, Daxx is able to regulate apoptosis, transcription, and cellular proliferation. Despite many studies into the function of Daxx, its precise role in the cell remains enigmatic. Herein, evidence is presented to expand upon the known anti-apoptotic function of Daxx, to establish Daxx as a novel molecular chaperone, and to further its repertoire of transcriptional targets. As an apoptotic inhibitor, Daxx is known to regulate p53 by stabilizing its main negative regulator, Mdm2, via formation of a ternary ...


Measuring Transcription Directly From Our Chromosomes, Marshall Levesque Jan 2013

Measuring Transcription Directly From Our Chromosomes, Marshall Levesque

Publicly Accessible Penn Dissertations

Our genome is organized into DNA segments called chromosomes. Alterations to the typically invariant number and composition of chromosomes are hallmarks of serious disease like cancer. Understanding how rearranging chromosomes affects chromosomal behavior and ultimately leads to disease requires chromosome-specific gene expression measurements, but current tools are insufficient. This thesis describes tools for measuring transcription while discriminating which copy of a gene the RNA comes from. The ability to take these measurements in single cells enabled us to measure changes in transcription on translocated chromosomes or from the maternal vs. paternal chromosomes.

Firstly, we introduce intron chromosomal expression FISH (iceFISH ...