Molecular Biology Commons^™

Dnp63a Suppresses Cell Invasion By Targeting Rac1 Through Mir-320a, Amjad Ahmed Aljagthmi

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DNp63a, a member of the p53 family of transcription factors, is overexpressed in a number of cancers and known to play a role in proliferation, differentiation, migration and invasion. DNp63a has been shown to regulate several microRNAs that play a role in both development and cancer, but to date there has not been a global analysis of p63- regulated miRNA. Using next-generation sequencing of small RNA from wild type and sip63 transfected HaCaT cells, our laboratory recently identified a number of DNp63a- regulated miRNAs by RNA-Seq studies which may serve as biomarkers of cancer progression. We identified a novel miRNA, …

Exploring The Role Of Lipin1 In Mitophagy Process Using Lipin1 Deficient-Egfp Tagged Lc3 Transgenic Mice, Abdullah Ali Alshudukhi

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Lipin1 (phosphatidic acid phosphatase) is a key molecule in the cells with two functions: first, it converts phosphatidic acid into diacylglycerol in the cytosol which in turn makes triglycerides. Second, in nucleus lipin1 acts as a transcriptional factor which regulates the expression of genes involved in the fatty acid oxidation and lipid metabolism. Clinically, Lpin1 gene mutations have been detected in patients with severe rhabdomyolysis accompanied with aggregated and dysfunctional mitochondria in their type II muscle fiber. Previously, we have observed that mice with lipin1 deficiency had aggregated mitochondria and abnormal autophagosomes formations by electron microscopy. The mechanism underlying the …

Investigation Of The Physiological Role Of Rin Gtpase In Cell Death, Axonal Injury, And Inflammation Following Traumatic Brain Injury, Megan Pannell

Theses and Dissertations--Molecular and Cellular Biochemistry

Traumatic brain injury (TBI) is a progressive disorder, in which the primary injury results in the initiation of a complex cascade of secondary biochemical and metabolic changes resulting in lasting neurological dysfunction and cognitive impairment. The heterogeneous nature of the disease has complicated the development of pharmacological agents to improve the outcomes of TBI; to date, no therapeutic treatment has been shown to be effective in clinical trials. Treatments targeting multiple secondary outcomes (cell death, axonal degeneration, and inflammation) may provide enhanced therapeutic efficacy following TBI.

Small Ras family GTP-binding proteins govern diverse cellular processes by directing the relay of …

Transcriptomic Analyses Of Cathatranthus Roseus Hairy Roots Overexpressing Crmyc2 And Orca3 And Roles Of Cross-Family Transcription Factor Interaction In Terpenoid Indole Alkaloid Biosynthesis, Xueyi Sui

Theses and Dissertations--Plant and Soil Sciences

Catharanthus roseus (Madagascar periwinkle), is a well-known medicinal plant that produces a vast array of terpenoid indole alkaloids (TIAs), including two anticancer compounds vinblastine and vincristine. Industrial scale production of TIAs is hampered by the difficulties of total chemical synthesis of these compounds and the fragmented knowledge on TIA pathway. Transcriptional regulation of the TIA biosynthetic pathway has not been thoroughly investigated in Catharanthus and only a few structural genes have been identified as the targets of two master regulators: the basic helix-loop-helix (bHLH) transcription factor (TF) CrMYC2 and APETALA2/ETHYLENE RESPONSE FACTOR (AP2/ERF), ORCA3. Next generation sequencing (NGS) has been …

Molecular Modeling Of Novel Tryptamine Analogs With Antibiotic Potential Through Their Inhibition Of Tryptophan Synthase, Jared Schattenkerk

CMC Senior Theses

The growing prevalence of antibiotic-resistant bacteria is a global health crisis that threatens the effectiveness of antibiotics in medical treatment. Increases in the number of antibiotic-resistant bacteria and a drop in the pharmaceutical development of novel antibiotics have combined to form a situation that is rapidly increasing the likelihood of a post-antibiotic era. The development of antibiotics with novel enzymatic targets is critical to stall this growing crisis. In silico methods of molecular modeling and drug design were utilized in the development of novel tryptamine analogs as potential antibiotics through their inhibition of the bacterial enzyme tryptophan synthase. Following the …

Development Of Molecular Diagnostic Tools For Mycobacterium Species, Hillary Bengtson

Electronic Theses and Dissertations

This dissertation focuses on the development of diagnostic tools for mycobacteria using hybridization based technologies including binary deoxyribozyme (BiDz) sensors and microarrays. The genus Mycobacterium, is a diverse group of bacteria containing 150+ species including M. tuberculosis (M.tb) and non-tuberculous mycobacteria (NTM) which exhibit a range of pathogenicity, drug susceptibility and growth characteristics. M. tuberculosis (M.tb) is the causative agent of tuberculosis (TB) and the leading cause of infectious disease related deaths worldwide. The control of TB is limited by the lack of sensitive and specific diagnostic tools available at the point of care (POC). The studies presented here illustrate …

The T6a Modification Acts As A Positive Determinant For The Anticodon Nuclease Prrc, And Is Distinctively Nonessential In Streptococcus Mutans, Jo Marie Bacusmo, Silvia S. Orsini, Jennifer Hu, Michael Demott, Patrick C. Thiaville, Ameer Elfarash, Mellie June Paulines, Diego Rojas-Benítez, Birthe Meineke, Christopher Deutsch, Dirk Iwata-Reuyl, Patrick A. Limbach, Peter C. Dedon, Kelly C. Rice, Stewart Shuman, Valérie De Crécy-Lagard

Chemistry Faculty Publications and Presentations

Endoribonuclease toxins (ribotoxins) are produced by bacteria and fungi to respond to stress, eliminate non-self competitor species, or interdict virus infection. PrrC is a bacterial ribotoxin that targets and cleaves tRNALys UUU in the anticodon loop. In vitro studies suggested that the post-transcriptional modification threonylcarbamoyl adenosine (t⁶A) is required for PrrC activity but this prediction had never been validated in vivo. Here, by using t⁶A-deficient yeast derivatives, it is shown that t⁶A is a positive determinant for PrrC proteins from various bacterial species. Streptococcus mutans is one of the few bacteria where the t …

Biochemical And Histological Differences Between Costal And Articular Cartilages, Michael W. Stacey

Bioelectrics Publications

Biologically, costal cartilage is an understudied tissue type and much is yet to be learned regarding underlying mechanisms related to form and function, and how these relate to disease states, specifically chest wall deformity. Chest wall deformities have a component of inheritance, implying underlying genetic causes; however the complexity of inheritance suggests multiple genetic components. At our Centre investigations were performed on gene expression of key select genes from costal cartilage removed at surgery of patients with chest wall deformity to show high expression of decorin, a key player in collagen fiber formation and growth. Also, the degree of tissue …

Clinicopathology And Molecular Determinants Underlying Benign Breast And Breast Cancer Lesions, Andreana Holowatyj Holowatyj

Wayne State University Dissertations

Despite converging incidence rates for breast cancers by race, disparities in mortality persist where black women suffer from poorer prognosis compared to white counterparts. To understand the clinical, demographic, and molecular characteristics underlying these disparities, we examined differences among patients with breast cancer to understand the role of human epidermal growth factor receptor 2 (HER2) status, age, and race/ethnicity among women diagnosed with hormone receptor-positive breast cancer, and disparities in surgical therapy among female patients with early stage young-onset breast cancer. Benign breast disease, another known risk factor for breast cancer, includes a histological spectrum of lesions, could contribute to …

Analysis Of The Secondary Neurodegenerative Consequences Of Primary Oligodendrocyte Stress Through The Use Of The Novel Obiden Mouse Model, Daniel Zdzislaw Radecki

Wayne State University Dissertations

The work of this project was to develop, test and characterize a potential novel mouse model of the neurodegenerative disease Multiple Sclerosis (MS). Historically, MS has been identified as a primary autoimmune disease of the central nervous system (CNS). However, treatments based on this view have met with limited success, and in most cases, fail to prevent progression of MS from mild to moderate and severe forms. Original observations regarding axonal and neuronal pathology in the white and gray matter of the CNS were rediscovered in the 1990s. These observations indicated that even in the absence of the immune system, …

Identification Of Oxygen Optima For Mouse Trophoblast Stem Cells And Human Embryos And The Stress Responses Upon Departing Optima, Yu Yang

Wayne State University Dissertations

Low level of oxygen (O2) occurs physiologically during in vivo embryo development. As developing embryos moving from fallopian tube to uterus, oxygen level gradually decreases to ≤ 5% at the time of blastocyst implantation. Blastocysts are made of two major cell populations, trophoblast cells and inner cell mass, from which trophoblast stem cells (TSCs) and embryonic stem cells (ESCs) are derived respectively. TSCs serve as placental stem cells that later on proliferate and differentiate into placenta. Previous study has shown that 2% O2 is the optimal O2 level for mTSC in vitro growth and potency maintenance, which agrees with their …

Effect Of Endoplasmic Reticulum Stress On Vascular Smooth Muscle Cells And Its Regulation Of Sm22Α, Neeraja Priyanka Annam

Wayne State University Dissertations

Background: The vascular smooth muscle cells(VSMC) possess the ability to differentiate into a synthetic phenotype in response to stress. This phenotypic modulation may be accompanied by inflammatory or osteogenic response in chronic stress. The synthetic state is characterized by low levels of contractile markers unlike the differentiated state.

Hypothesis: Endoplasmic reticulum (ER) stress causes phenotypic modulation in VSMCs leading to apoptosis. Many transcription factors induced by ER stress contribute to the downregulation of Sm22α. Perturbation in cytoskeletal dynamics exacerbates the ER stress response.

Methods: Ex-vivo culture was used to establish importance of Sm22 in ER stress. In vitro analysis was …

Sh3 And Multiple Ankyrin Repeat Domain 3 (Shank3) Affects The Expression Of Hyperpolarization-Activated Cyclic Nucleotide-Gated (Hcn) Channels In Mouse Models Of Autism, Nikhil N. Shah

Theses and Dissertations

SH3 and multiple ankyrin repeat domains 3 (SHANK3) is a multidomain scaffold protein that is highly augmented in the postsynaptic density (PSD) of excitatory glutamatergic synapses within the central and peripheral nervous systems. SHANK3 links neurotransmitter receptors, ion channels, and other critical membrane proteins to intracellular cytoskeleton and signal transduction pathways. Mutations in SHANK3 are linked with a number neuropsychiatric disorders including autism spectrum disorders (ASDs). Intellectual disability, impaired memory and learning, and epilepsy are some of the deficits commonly associated with ASDs that result from mutations in SHANK3. Interestingly, these symptoms show some clinical overlap with presentations of human …

Epigenetic Editing To Validate Findings From Methylome-Wide Association Studies Of Neuropsychiatric Disorders, Robin F. Chan

Theses and Dissertations

DNA methylation is necessary for learning, memory consolidation and has been implicated in a number of neuropsychiatric disorders. Obtaining high quality and comprehensive data for the three common forms of methylation in brain is challenging for methylome-wide association studies (MWAS). To address this we optimized a panel of enrichment methods for screening the brain methylome. Results show that these enrichment techniques approach the coverage and fidelity of the current gold standard bisulfite based techniques. Our MBD-based method can also be used with low amounts of genomic material from limited human biomaterials. Psychiatric disorders have high prevalence and are often chronic …

Relb Acts As A Molecular Switch To Drive Chronic Inflammation In Glioblastoma Multiforme (Gbm)., Michael R. Waters

Theses and Dissertations

Inflammation is a homeostatic response to tissue injury or infection, which is normally short- lived and quickly resolves to limit tissue damage. In contrast, chronic inflammation has been linked to a variety of human diseases, including cancers such as glioblastoma multiforme (GBM). GBMs are very aggressive tumors with very low patient survival rates, which have not improved in several decades. GBM tumors are characterized by necrosis and profound inflammation; with cytokines secreted by both GBM cells and the tumor microenvironment. The mechanisms by which chronic inflammation develops and persists in GBM regardless of multiple anti-inflammatory feedback loops remain elusive. This …

Illuminate The Pathway Of Membrane Protein Association And Degradation, Zhaoshuai Wang

Theses and Dissertations--Chemistry

Escherichia coli transporter protein AcrB and its homologues are the inner membrane components of the Resistance-Nodulation-Division (RND) family efflux pumps in Gram-negative bacteria. It is well accepted that soluble proteins are only marginally stable, but such insight is missing for membrane proteins. The lack of stability data, including thermodynamic stability and oligomer association affinity is a result of intrinsic difficulties in working with membrane proteins. In addition, the degradation of soluble proteins in E. coli has been extensively studied whereas the degradation process of membrane proteins remains unclear. A focus of my thesis is the validation and development of methods …

High-Throughput Single-Molecule Mapping Links Subtelomeric Variants And Long-Range Haplotypes With Specific Telomeres, Eleanor Young, Steven Pastor, Ramakrishnan Rajagopalan, Jennifer Mccaffrey, Justin Sibert, Angel C. Y. Mak, Pui-Yan Kwok, Harold Riethman, Ming Xiao

School of Medical Diagnostics & Translational Sciences Publications

Accurate maps and DNA sequences for human subtelomere regions, along with detailed knowledge of subtelomere variation and long-range telomereterminal haplotypes in individuals, are critical for understanding telomere function and its roles in human biology. Here, we use a highly automated whole genome mapping technology in nano-channel arrays to analyze large terminal human chromosome segments extending from chromosome-specific subtelomere sequences through subtelomeric repeat regions to terminal (TTAGGG) n repeat tracts. We establish detailed maps for subtelomere gap regions in the human reference sequence, detect many new large subtelomeric variants and demonstrate the feasibility of long-range haplotyping through segmentally duplicated subtelomere regions. …

Tissue Specific Microenvironments: A Key Tool For Tissue Engineering And Regenerative Medicine, Patrick C. Sachs, Peter A. Mollica, Robert D. Bruno

School of Medical Diagnostics & Translational Sciences Publications

The accumulated evidence points to the microenvironment as the primary mediator of cellular fate determination. Comprised of parenchymal cells, stromal cells, structural extracellular matrix proteins, and signaling molecules, the microenvironment is a complex and synergistic edifice that varies tissue to tissue. Furthermore, it has become increasingly clear that the microenvironment plays crucial roles in the establishment and progression of diseases such as cardiovascular disease, neurodegeneration, cancer, and ageing. Here we review the historical perspectives on the microenvironment, and how it has directed current explorations in tissue engineering. By thoroughly understanding the role of the microenvironment, we can begin to correctly …

Potential Efficacy Of Targeting Mll1 In Breast Cancer, Austin Gable Holmes

Master's Theses

In 2017, it is estimated that breast cancer will be the most prevalent newly diagnosed cancer in females, at 30% in the U.S. alone1. From the early 1990s to 2014 the death rate in females from breast cancer has dropped by 10% but still stands as the second highest cancer related death in females in the U.S.2. One of the biggest hurdles in breast cancer research is disease heterogeneity. New mechanisms of disease development and progression are encountered frequently. One mechanism studied in the past decades is epigenetics. It has been theorized that the cancer epigenome can maintain an abnormal …

Evidence Of A Perilipin 5 Splice Variant, Brodie Ranzau

Undergraduate Honors Thesis Projects

Alternative splicing occurs throughout the human genome, leading to multiple proteins from a single gene. The resulting proteins can be nearly identical or vastly different in how they function. Alternative splicing within the perilipin family has been observed in perilipins 1 and 3, giving rise to proteins with varying functions. In the course of our studies on perilipin 5, an immunoblot signal was observed that corresponded to an approximately 35 kDa protein. Western blot analysis of this protein has revealed its expression in C2C12 cultured myoblasts and in heart and liver mouse tissue. Further analysis of perilipin 5 cDNA through …

Navigating Human Cytomegalovirus (Hcmv) Envelopment And Egress, William Longeway Close

Wayne State University Dissertations

Human cytomegalovirus (HCMV) is a ubiquitous viral pathogen. In individuals with fully functioning and mature immune systems, HCMV is associated with mild symptoms prior to establishing latency. In individuals with naïve or compromised immune systems, HCMV is capable of causing severe organ damage. HCMV is the leading infectious cause of congenital birth defects and a major non-genetic cause of hearing loss. Unfortunately, antiviral treatment options lack diversity due to limited knowledge of virion replication. If HCMV replication were better understood, new antiviral treatments could be developed.

In this work, we describe the development and implementation of new tools to study …

Expression, Purification And Characterization Of Lysine Methyltransferase Smyd5, Wen Xue

Wayne State University Theses

Methylation of histones and non-histone proteins play vital roles in numerous cellular processes including gene expression regulation and DNA damage response. The identifications of methyltransferase SMYD protein family are not well characterized. SMYD5 is a unique and critical member of SMYD family involved in immune response, stem cell renew, hematopoiesis regulation and cancer metastasis. Understanding its function and structure is monumental to human disease. With the achievement of SMYD5 expression and purification, the association between SMYD5 and its substrate histone H4 has been investigated. While possessing a poly-E tail instead of the TPR domain included in other members, SMYD5 is …

Biochemical Analysis Of Putative Single-Stranded Nucleic Acid Binding Proteins In Porphyromonas Gingivalis, Steve H. Kokorelis

Theses and Dissertations

Proteins that bind to both DNA and RNA embody the ability to perform multiple functions by a single gene product. These nucleic acid binding proteins in prokaryotes can play a vital role in many cellular processes, including replication, transcription, gene expression, recombination, and repair, to name a few. Nucleic acid binding proteins have unique functional characteristics that stem from their structural attributes that have evolved in a widely-conserved manner. In Escherichia coli (E. coli), the highly-conserved histone-like protein, HU, which predominates as a heterodimer of HUα and HUβ, has been found to bind to both dsDNA and ssDNA. …

An Effective Computational Method Incorporating Multiple Secondary Structure Predictions In Topology Determination For Cryo-Em Images, Abhishek Biswas, Desh Ranjan, Mohammad Zubair, Stephanie Zeil, Kamal Al Nasr, Jing He

Computer Science Faculty Publications

A key idea in de novo modeling of a medium-resolution density image obtained from cryo-electron microscopy is to compute the optimal mapping between the secondary structure traces observed in the density image and those predicted on the protein sequence. When secondary structures are not determined precisely, either from the image or from the amino acid sequence of the protein, the computational problem becomes more complex. We present an efficient method that addresses the secondary structure placement problem in presence of multiple secondary structure predictions and computes the optimal mapping. We tested the method using 12 simulated images from alpha-proteins and …

Transcriptional Regulation Of Skeletal Muscle Atrophy-Induced Gene Expression By Muscle Ring Finger-1 And Myogenic Regulatory Factors, Karina Marie Kakareka

UNF Graduate Theses and Dissertations

Skeletal muscle wasting occurs as a corollary of numerous physiological conditions, including denervation, immobilization, and aging. The E3 ubiquitin ligases MuRF1 and MAFbx are induced under nearly all atrophy conditions and are believed to play a key role in protein degradation. Data in this thesis provides interesting new evidence that MuRF1 may also act as a transcriptional modulator of atrophy-induced genes or atrogenes. The transcriptional regulation of MuRF1 and MAFbx were characterized using a reporter gene system and exhibited repressed activity in C₂C₁₂ cells overexpressing MuRF1. Furthermore, ectopic expression of the myogenic regulatory factors (MRFs), MyoD and …

A Rare Case Of Erythema Elevatum Diutinum Presenting As Diffuse Neuropathy, G H. Nguyen, E L. Guo, D Norris

Faculty, Staff and Students Publications

No abstract provided.

Disrupted Superior Collicular Activity May Reveal Cervical Dystonia Disease Pathomechanisms, Eavan M. Mcgovern, Owen Killian, Shruti Narashimham, Brendan Quinlivan, John Butler, Rebecca Beck, Ines Beiser, Laura W. Williams, Ronan P. Killeen, Michael Farrell, Sean O'Riordan, Richard B. Reilly, Michael Hutchinson

Articles

Cervical dystonia is a common neurological movement disorder characterised by muscle contractions causing abnormal movements and postures affecting the head and neck. The neural networks underpinning this condition are incompletely understood. While animal models suggest a role for the superior colliculus in its pathophysiology, this link has yet to be established in humans. The present experiment was designed to test the hypothesis that disrupted superior collicular processing is evident in affected patients and in relatives harbouring a disease-specific endophenotype (abnormal temporal discrimination). The study participants were 16 cervical dystonia patients, 16 unaffected first-degree relatives with abnormal temporal discrimination, 16 unaffected …

Characterization Of A Putative Hemolysin Expressed By Sneathia Amnii, A Preterm Birth-Associated Pathogen, Lizette Carrasco, Kimberly Jefferson

Undergraduate Research Posters

The gram-negative bacteria Sneathia amnii is a poorly-characterized commensal of the female urogenital tract frequently associated with adverse clinical outcomes such as bacterial vaginosis (BV), amnionitis, and preterm labor. To investigate its potential role in virulence, we sought to identify and characterize virulence determinants produced by S. amnii in an effort to better understand the pathogenesis of infectious preterm birth. Through sequencing of the Sn35 genome (type strain of S. amnii), we identified two genes with amino acid sequence similarity and structural similarity to the filamentous hemagglutinin (FHA) protein of Bordetella pertussis and its Type Vb transporter. Because S. amnii …

Separation Of Blood Mixtures Using Fluorescently Labeled Antibodies, Dani Jabado

Undergraduate Research Posters

Identifying and analyzing biological mixture samples at a crime scene are of paramount concern for forensic scientists, especially if that type of evidence contains only one cell type. The presence of multiple contributors in a biological evidence sample reduces the probative value of DNA evidence and can sometimes lead to its eventual loss of value. As such, this study was performed in an attempt to examine and evaluate flow cytometry analysis as a means to separate blood mixture samples labeled with fluorescent antibodies. Fluorescein Isothiocyanate (FITC) antibodies were specifically targeted and bound to HLA (Human Leukocyte Antigens) markers present on …

Evolution Of Protein Complexes In Bacterial Species, Shwetha Hara Sridhar, Wedad Albalawi, Peter Uetz

Undergraduate Research Posters

Protein complexes are composed of two or more associated polypeptide chains that may have different functions. Protein complexes play a critical role for all processes in life and are considered as highly conserved in evolution. In previous studies, protein complexes from E. coli or Mycoplasma pneumoniae have been characterized experimentally, revealing that a typical bacterial cell has on the order of 500 protein complexes. Using gene homology (orthology), these experimentally-observed complexes can be used to predict protein complexes across many species of bacteria. Surprisingly, the majority of protein complexes is not conserved, demonstrating an unexpected evolutionary flexibility.

The current research …