Genetic Structures Commons^™

Characterization Of The Overexpression Of Reca Homologs Rad51 And Dmc1 In Tetrahymena Thermophila, Jianna M. Cox

Graduate Theses/Dissertations

RecA homologs, Dmc1 and Rad51, work to repair DNA double-strand breaks (DSBs) within the cell through the recombination of homologous sections of DNA. Dmc1 works to repair programmed DSBs through meiotic recombination, while Rad51 functions to repair both meiotic and non-meiotic DSBs, the latter repaired through the process of homologous recombination repair (HHR). Chemotherapeutics, exogenous agents, work to form DSBs in cancer cells, attempting to inhibit the cell’s growth. A hyper recombinant phenotype is often seen in cancer cells due to the overexpression of RAD51, leading to drug resistance, the persistence of cancers, and an overall poor patient outcome. …

Long G4-Rich Enhancers Physically Interacts With Promoters Via A G4:G4 Dna-Based Mechanism, Jeffrey David Demeis

Graduate Theses and Dissertations (2019 - present)

Enhancers are genomic sequences that function as regulatory elements capable of increasing the transcription of a given gene often located at a considerable distance. The broadly accepted model of enhancer activation involves bringing an enhancer-bound activator protein complex into close spatial proximity to its target promoter through chromatin looping. Equally relevant to the work described herein, roles for guanine (G) rich sequences in transcriptional regulation are now widely accepted. Non-coding G-rich sequences are commonly found in gene promoters and enhancers, and various studies have described specific instances where G-rich sequences regulate gene expression via their capacity to form G-quadruplex (G4) …

Setd2 Safeguards The Genome Against Isochromosome Formation, Frank M Mason, Emily S Kounlavong, Anteneh T Tebeje, Rashmi Dahiya, Tiffany Guess, Abid Khan, Logan Vlach, Stephen R Norris, Courtney A Lovejoy, Ruhee Dere, Brian D Strahl, Ryoma Ohi, Peter Ly, Cheryl Lyn Walker, W Kimryn Rathmell

Faculty, Staff and Students Publications

Isochromosomes are mirror-imaged chromosomes with simultaneous duplication and deletion of genetic material which may contain two centromeres to create isodicentric chromosomes. Although isochromosomes commonly occur in cancer and developmental disorders and promote genome instability, mechanisms that prevent isochromosomes are not well understood. We show here that the tumor suppressor and methyltransferase SETD2 is essential to prevent these errors. Using cellular and cytogenetic approaches, we demonstrate that loss of SETD2 or its epigenetic mark, histone H3 lysine 36 trimethylation (H3K36me3), results in the formation of isochromosomes as well as isodicentric and acentric chromosomes. These defects arise during DNA replication and are …

Decidualization Of Human Endometrial Stromal Cells Requires Steroid Receptor Coactivator-3, Vineet K Maurya, Maria M Szwarc, David M Lonard, William E Gibbons, San-Pin Wu, Bert W O'Malley, Francesco J Demayo, John P Lydon

Faculty, Staff and Students Publications

Steroid receptor coactivator-3 (SRC-3; also known as NCOA3 or AIB1) is a member of the multifunctional p160/SRC family of coactivators, which also includes SRC-1 and SRC-2. Clinical and cell-based studies as well as investigations on mice have demonstrated pivotal roles for each SRC in numerous physiological and pathophysiological contexts, underscoring their functional pleiotropy. We previously demonstrated the critical involvement of SRC-2 in murine embryo implantation as well as in human endometrial stromal cell (HESC) decidualization, a cellular transformation process required for trophoblast invasion and ultimately placentation. We show here that, like SRC-2, SRC-3 is expressed in the epithelial and stromal …

Identifying The Molecular Cause Of Extreme Endoplasmic Reticulum Dilation In Pediatric Osteosarcoma And Its Relationship To The Disease, Rachael Wood

Theses and Dissertations (ETD)

Pediatric osteosarcoma tumors are characterized by an unusual abundance of grossly dilated endoplasmic reticulum and an immense genomic instability that has complicated identifying new effective molecular therapeutic targets. Here we report a novel molecular signature that encompasses the majority of 108 patient tumor samples, PDXs and osteosarcoma cell lines. These tumors exhibit reduced expression of four critical COPII vesicle proteins that has resulted in the accumulation of procollagen-I protein within ‘hallmark’ dilated ER. Using CRISPR activation technology, increased expression of only SAR1A and SEC24D to physiologically normal levels was sufficient to restore both collagen-I secretion and resolve dilated ER morphology …

Suppression Of Human Coronavirus 229e Infection In Lung Fibroblast Cells Via Rna Interference, Hamidreza Montazeri Aliabadi, Jennifer Totonchy, Parvin Mahdipoor, Keykavous Parang, Hasan Uludağ

Pharmacy Faculty Articles and Research

Despite extensive efforts to repurpose approved drugs, discover new small molecules, and develop vaccines, COVID-19 pandemic is still claiming victims around the world. The current arsenal of antiviral compounds did not perform well in the past viral infections (e.g., SARS), which casts a shadow of doubt for use against the new SARS-CoV-2. Vaccines should offer the ultimate protection; however, there is limited information about the longevity of the generated immunity and the protection against possible mutations. This study uses Human Coronavirus 229E as a model coronavirus to test the hypothesis that effective delivery of virus-specific siRNAs to infected cells will …

Discovery And Characterization Of Bromodomain 2-Specific Inhibitors Of Brdt, Zhifeng Yu, Angela F Ku, Justin L Anglin, Rajesh Sharma, Melek Nihan Ucisik, John C Faver, Feng Li, Pranavanand Nyshadham, Nicholas Simmons, Kiran L Sharma, Sureshbabu Nagarajan, Kevin Riehle, Gundeep Kaur, Banumathi Sankaran, Marta Storl-Desmond, Stephen S Palmer, Damian W Young, Choel Kim, Martin M Matzuk

Faculty, Staff and Students Publications

Bromodomain testis (BRDT), a member of the bromodomain and extraterminal (BET) subfamily that includes the cancer targets BRD2, BRD3, and BRD4, is a validated contraceptive target. All BET subfamily members have two tandem bromodomains (BD1 and BD2). Knockout mice lacking BRDT-BD1 or both bromodomains are infertile. Treatment of mice with JQ1, a BET BD1/BD2 nonselective inhibitor with the highest affinity for BRD4, disrupts spermatogenesis and reduces sperm number and motility. To assess the contribution of each BRDT bromodomain, we screened our collection of DNA-encoded chemical libraries for BRDT-BD1 and BRDT-BD2 binders. High-enrichment hits were identified and resynthesized off-DNA and examined …

Investigating The Role Of Znf384 Rearrangements In Acute Leukemia, Kirsten Dickerson

Theses and Dissertations (ETD)

Chromosomal rearrangements involving ZNF384 are the defining lesion in 5% of pediatric and adult B-cell acute lymphoblastic leukemia and tumors are characterized by aberrant myeloid marker expression. Additionally, ZNF384 rearrangements are the defining lesion in nearly half of pediatric B/myeloid mixed phenotype acute leukemia. These fusions juxtapose full-length ZNF384 to the N terminal portion of a diverse range of partners, most often, transcription factors or epigenetic modifiers. It has been shown that ZNF384-rearranged tumors have a distinct gene expression profile that is consistent between disease groups and N terminal partners. Genomic analyses of patient tumors has shown that ZNF384 fusions …

Yeast Mitochondrial Protein Pet111p Binds Directly To Two Distinct Targets In Cox2 Mrna, Suggesting A Mechanism Of Translational Activation, Julia L Jones, Katharina B Hofmann, Andrew T Cowan, Dmitry Temiakov, Patrick Cramer, Michael Anikin

Rowan-Virtua School of Osteopathic Medicine Departmental Research

The genes in mitochondrial DNA code for essential subunits of the respiratory chain complexes. In yeast, expression of mitochondrial genes is controlled by a group of gene-specific translational activators encoded in the nucleus. These factors appear to be part of a regulatory system that enables concerted expression of the necessary genes from both nuclear and mitochondrial genomes to produce functional respiratory complexes. Many of the translational activators are believed to act on the 5'-untranslated regions of target mRNAs, but the molecular mechanisms involved in this regulation remain obscure. In this study, we used a combination of in vivo and in …

N-Terminal Domain Of Human Uracil Dna Glycosylase (Hung2) Promotes Targeting To Uracil Sites Adjacent To Ssdna-Dsdna Junctions, Brian P Weiser, Gaddiel Rodriguez, Philip A Cole, James T Stivers

Rowan-Virtua School of Osteopathic Medicine Departmental Research

The N-terminal domain (NTD) of nuclear human uracil DNA glycosylase (hUNG2) assists in targeting hUNG2 to replication forks through specific interactions with replication protein A (RPA). Here, we explored hUNG2 activity in the presence and absence of RPA using substrates with ssDNA-dsDNA junctions that mimic structural features of the replication fork and transcriptional R-loops. We find that when RPA is tightly bound to the ssDNA overhang of junction DNA substrates, base excision by hUNG2 is strongly biased toward uracils located 21 bp or less from the ssDNA-dsDNA junction. In the absence of RPA, hUNG2 still showed an 8-fold excision bias …

Mechanism Of Transcription Anti-Termination In Human Mitochondria., Hauke S Hillen, Andrey V Parshin, Karen Agaronyan, Yaroslav I Morozov, James J Graber, Aleksandar Chernev, Kathrin Schwinghammer, Henning Urlaub, Michael Anikin, Patrick Cramer, Dmitry Temiakov

Rowan-Virtua School of Osteopathic Medicine Departmental Research

In human mitochondria, transcription termination events at a G-quadruplex region near the replication origin are thought to drive replication of mtDNA by generation of an RNA primer. This process is suppressed by a key regulator of mtDNA-the transcription factor TEFM. We determined the structure of an anti-termination complex in which TEFM is bound to transcribing mtRNAP. The structure reveals interactions of the dimeric pseudonuclease core of TEFM with mobile structural elements in mtRNAP and the nucleic acid components of the elongation complex (EC). Binding of TEFM to the DNA forms a downstream "sliding clamp," providing high processivity to the EC. …

Endonucleolytic Cleavage In The Expansion Segment 7 Of 25s Rrna Is An Early Marker Of Low-Level Oxidative Stress In Yeast, Daniel Shedlovskiy, Jessica A Zinskie, Ethan Gardner, Dimitri G Pestov, Natalia Shcherbik

Rowan-Virtua School of Osteopathic Medicine Departmental Research

The ability to detect and respond to oxidative stress is crucial to the survival of living organisms. In cells, sensing of increased levels of reactive oxygen species (ROS) activates many defensive mechanisms that limit or repair damage to cell components. The ROS-signaling responses necessary for cell survival under oxidative stress conditions remain incompletely understood, especially for the translational machinery. Here, we found that drug treatments or a genetic deficiency in the thioredoxin system that increase levels of endogenous hydrogen peroxide in the yeast Saccharomyces cerevisiae promote site-specific endonucleolytic cleavage in 25S ribosomal RNA (rRNA) adjacent to the c loop of …

Il-15 Mediates Mitochondrial Activity Through A Ppar𝛿-Dependent-Ppar𝛼-Independent Mechanism In Skeletal Muscle Cells, Shantaé M. Thornton, James E. Krolopp, Marcia J. Abbott

Health Sciences and Kinesiology Faculty Articles

Molecular mediators of metabolic processes, to increase energy expenditure, have become a focus for therapies of obesity. The discovery of cytokines secreted from the skeletal muscle (SKM), termed “myokines,” has garnered attention due to their positive effects on metabolic processes. Interleukin-15 (IL-15) is a myokine that has numerous positive metabolic effects and is linked to the PPAR family of mitochondrial regulators. Here, we aimed to determine the importance of PPAR𝛼 and/or PPAR𝛿 as targets of IL-15 signaling. C2C12 SKM cells were differentiated for 6 days and treated every other day with IL-15 (100 ng/mL), a PPAR𝛼 inhibitor (GW-6471), a PPAR𝛿 …

Investigation Of Ultrasound Targeted Microbubbles As A Therapeutic Gene Delivery System For Prostate Cancer, Rounak Paramjeet Nande

Theses, Dissertations and Capstones

A major challenge for effective gene therapy is systemic delivery of viruses carrying therapeutic genes into affected tissue. The immunogenic nature of human adenoviruses (Ads) limits their use for intratumoral (IT) injection in gene therapy. Ads transfection is further hampered by the fluctuating presence of Coxsackie and Adenovirus Receptor (CAR) and integrins on the cells’ surface. To circumvent these limitations we developed a novel approach wherein Ads are encapsulated inside the shell of lyophilized, lipid-encapsulated, perfluorocarbon microbubbles (MBs)/ultrasound (US) contrast agents, which act as delivery vehicles for a sitespecific gene transfer system.

We performed infection studies with Ad.GFP (Green Fluorescent …

Lineage-Specific Interface Proteins Match Up The Cell Cycle And Differentiation In Embryo Stem Cells, Angela Re, Christopher T. Workman, Levi Waldron, Alessandro Quattrone, Søren Brunak

Publications and Research

The shortage of molecular information on cell cycle changes along embryonic stem cell (ESC) differentiation prompts an in silico approach, which may provide a novel way to identify candidate genes or mechanisms acting in coordinating the two programs. We analyzed germ layer specific gene expression changes during the cell cycle and ESC differentiation by combining four human cell cycle transcriptome profiles with thirteen in vitro human ESC differentiation studies. To detect cross-talk mechanisms we then integrated the transcriptome data that displayed differential regulation with protein interaction data. A new class of non-transcriptionally regulated genes was identified, encoding proteins which interact …

Role Of Iron In Ethanol Derived Hepatic Stress, Jesse A. Thornton

Theses, Dissertations and Capstones

Chronic alcohol abuse is the third leading cause of preventable death in the United States. Ethanol metabolism causes liver injury through alterations in hepatic metabolic state, redox status, and acetaldehyde adduct formations. Increased iron absorption is associated with chronic ethanol consumption and may play a role in ethanol induced oxidative stress. We tested the hypothesis that normal labile iron in the liver plays a role in ethanol related pathological stress, using C57/Bl6 mice pair-fed Lieber-DeCarli liquid ethanol diets for 11 and 22 weeks. Normal iron group mice received 55mg/kg iron as ferric citrate, whereas the low iron groups received 5mg/kg. …

Regulation Of Biofilm Formation Of Pseudomonas Aeruginosa, Nathaniel Edwards Head

Theses, Dissertations and Capstones

Cystic fibrosis (CF) is the most common, autosomal recessive lethal genetic disease in the Caucasian population, resulting from a malfunctioned cystic fibrosis transmembrane conductance regulator (CFTR) and leading to bacterial lung infections. P. aeruginosa, an opportunistic pathogen, establishes a chronic infection in CF with a phenotype of overproduction of an exopolysaccharide (alginate) due to host-directed mutagenesis. While free-floating planktonic bacteria can be properly cleared from the CF lung, P. aeruginosa, along with alginate production, establishes an infection in the form of a biofilm which supports its survival in nature and in vivo. As a result, genomic structure, …

Phosphorylation Of The Rex Protein Of Human T-Cell Leukemia Virus Type 1, Yoshifumi Adachi, Terry D. Copeland, Chiaki Takahashi, Tetsuya Nosaka, Aftab Ahmed, Stephen Oroszlan, Masakazu Hatanaka

Pharmacy Faculty Articles and Research

Rex protein, the posttranscriptional regulator of human T-cell leukemia virus type I (HTLV-I), is required for the control of viral structural protein expression and virus replication. Rex is a phosphoprotein found predominantly in the cell nucleolus, whose function is thought to be regulated by its nucleolar localization and phosphorylation. Therefore, we investigated the in vivo phosphorylation of Rex protein in more detail. Phosphorylation of Rex occurred in all HTLV-I-infected cell lines examined in vivo, primarily at serine residues and to a very small extent at threonine residues. Treatment of cells with 12-O-tetradecanoylphorbol-13-acetate (TPA) led to significant but transient enhancement of …