Molecular Biology Commons^™

The Sex Of The Cell Dictates Its Response, Carlos Ganesh Penaloza

Dissertations, Theses, and Capstone Projects

Male and female differences in frequency of occurrence in disease have perplexed scientists for some time. This in part derives from limitations in the systems in which one can evaluate sex differences. At the organismal level, differences can be hidden by a myriad of extensive and complex factors. Additional limitations exist since most biomedical studies are performed almost exclusively on male subjects, as the female hormonal milieu is intrinsically more variable and too troublesome for routine inclusion in research protocols. Research documenting sex differences continues to grow, and while most researchers suggests that sex hormones are at the core of …

Strong Dna Deformation Required For Extremely Slow Dna Threading Intercalation By A Binuclear Ruthenium Complex, Ali A. Almaqwashi, Thayaparan Paramanathan, Per Lincoln, Ioulia Rouzina, Fredrik Westerlund, Mark C. Williams

Physics Faculty Publications

DNA intercalation by threading is expected to yield high affinity and slow dissociation, properties desirable for DNA-targeted therapeutics. To measure these properties, we utilize single molecule DNA stretching to quantify both the binding affinity and the force-dependent threading intercalation kinetics of the binuclear ruthenium complex Δ,Δ-[μ‐bidppz‐(phen)₄Ru₂]⁴⁺ (Δ,Δ-P). We measure the DNA elongation at a range of constant stretching forces using optical tweezers, allowing direct characterization of the intercalation kinetics as well as the amount intercalated at equilibrium. Higher forces exponentially facilitate the intercalative binding, leading to a profound decrease in the binding site size that …

The Effects Of Lipoxin A4 (Lxa4) On Neutrophil Biology In Sepsis, Benedict Wu

Graduate School of Biomedical Sciences Theses and Dissertations

During sepsis, neutrophils are inappropriately activated and have prolonged lifespans, thus becoming dysfunctional. Excessive neutrophil activation can lead to tissue injury while neutrophil dysfunction can lead to decreased free radical production and reduced phagocytosis, preventing the host from clearing preexisting infections. Lipoxin A4 (LXA4) is a specialized pro-resolution mediator which reduces neutrophil migration and expression of proinflammatory mediators. Intact neutrophil functions are critical for the host to efficiently clear invading pathogens. The effects of LXA4 on neutrophil function in sepsis have not been established. Using the cecal ligation and puncture (CLP) model of sepsis, LXA4 administered 1 h after sepsis …

Preliminary Study Of Kleptoplasty In Foraminifera Of South Carolina, Shawnee Lechliter

Bridges: A Journal of Student Research

Recent studies of living foraminifera, microscopic aquatic protists, indicate that some species have the ability to steal photosynthetic plastids from other microorganism and keep them viable through a process called kleptoplasty. Studying the symbiotic relationships within these diverse protists gives insight not only into evolutionary history, but also their importance to the ecosystem. We determined the presence of these kleptoplastic species and identified presence and origin of sequestered plastids based on morphological identification and molecular data from samples collected at Waties Island, South Carolina. We identified two kleptoplastic genera (Elphidium and Haynesina) and two non-kleptoplastic genera (Ammonia and Quinqueloculina) present …

The Role Of Mammalian Cyclin C In Stress Response And Thyroid Disease Progression, Kun Wang

Graduate School of Biomedical Sciences Theses and Dissertations

The cellular response to stress involves changes in gene expression programs as well as the remodeling of subcellular organelles including the mitochondria. Important for this study, the mitochondria undergo extensive fragmentation when the cell is exposed to a variety of stressors. The cyclin C-Cdk8 kinase is a highly conserved transcription factor that associates with the RNA polymerase II holoenzyme. Using cyclin C knockout (CCNC^-/-) mouse embryonic fibroblast (MEF) cells, I revealed that cyclin C is required for the extensive mitochondrial fragmentation following treatment with pro-oxidants or the anti-cancer drug cisplatin. However, Cdk8 is not required for this process …

Formyl Group Activation Of Bromopyrrole Suzuki Cross-Coupling: Application To A Formal Synthesis Of Lamellarin G Trimethyl Ether, Andrew Harrison

Honors Theses

Many biologically interesting compounds have been isolated from marine natural products, many of which contain characteristic pyrroles. Compounds such as polycitones, storniamides, ningalins, and lamellarins have been of particular interest for synthesis due to their vast pharmaceutical potential including the ability to fight tumors as well as induce cytolysis of drug resistant cancer cell lines. Previous studies on the synthesis of Lamellarin G trimethyl ether by the Gupton Group have relied upon vinylogous amide derivatives as building blocks.2 A new pathway utilizing a formylated pyrrole ring provides an interesting method for the synthesis of Lamellarin G trimethyl ether. Regioselective Suzuki …

Regulation Of Nkcc2 Trafficking By Vesicle Fusion Proteins Vamp2 And Vamp3 In The Thick Ascending Limb, Paulo Sebastian Caceres Puzzella

Wayne State University Dissertations

The thick ascending limb (TAL) in the kidney regulates extracellular fluid volume and blood pressure. The Na/K/2Cl cotransporter NKCC2 plays a central role in NaCl absorption by the TAL and blood pressure. NKCC2 trafficking to the apical membrane is a major mechanism to control NKCC2 activity. However, little is known about the proteins that mediate NKCC2 trafficking. Inhibition of the vesicle fusion proteins VAMP2 and VAMP3 blunts the increase in surface NKCC2 expression and NaCl absorption in response to stimulation by cAMP. In other cells, VAMPs mediate fusion of exocytic vesicles with the plasma membrane. Whether VAMP2 and VAMP3 mediate …

The Mechanism Of Regulation Of Autosomal Heterochromatic Genes In Drosophila Melanogaster Males By Rox Rna And Msl Proteins, Satya Kiran Koya

Wayne State University Dissertations

In humans and flies, males and females have different set of sex chromosomes contributing to different levels of X-linked gene expression. To equalize X-linked gene dosage between sexes, both humans and flies developed independent strategies which are called dosage compensation. Human females randomly inactivate one of their X chromosome into barr body and Drosophila males up regulate their single X chromosome two fold. Both strategies equalize of X linked gene dose between sexes.

In Drosophila, dosage compensation is brought about by the ribonucleoprotein Male Specific Lethal (MSL) complex that binds hundreds of sites along the X chromosome and modifies …

Nuclear Import And Interactions Of Potato Yellow Dwarf Virus Nucleocapsid, Matrix, And Phosphoprotein, Gavin Lloyd Franklin Anderson

Theses and Dissertations--Plant Pathology

Potato yellow dwarf virus (PYDV) is the type species of the genus Nucleorhabdovirus and, like all members of this genus, replication and morphogenesis occurs inside the nuclei of infected cells. Protein localization prediction algorithms failed to identify a nuclear localization signal (NLS) in PYDV nucleocapsid (N) protein, although PYDV-N has been shown to localize exclusively to the nucleus when expressed as a green fluorescent protein (GFP):N fusion in plant cells. Deletion analysis and alanine-scanning mutagenesis identified two amino acid motifs, ⁴¹⁹QKR⁴²¹ and ⁴³²KR⁴³³, that were shown to be essential for nuclear import and interaction with …

Ether Bridge Formation And Chemical Diversification In Loline Alkaloid Biosynthesis, Juan Pan

Theses and Dissertations--Plant Pathology

Loline alkaloids, found in many grass-Epichloë symbiota, are toxic or feeding deterrent to invertebrates. The loline alkaloids all share a saturated pyrrolizidine ring with a 1-amine group and an ether bridge linking C2 and C7. The steps in biosynthesis of loline alkaloids are catalyzed by enzymes encoded by a gene cluster, designated LOL, in the Epichloë genome. This dissertation addresses the enzymatic, genetic and evolutionary basis for diversification of these alkaloids, focusing on ether bridge formation and the subsequent modifications of the 1-amine to form different loline alkaloids.

Through gene complementation of a natural lolO mutant and comparison …

A Novel Function For 12-Lipoxygenase In C-Met And Integrin Β4 Axis Crosstalk, Elizabeth Tovar

Wayne State University Dissertations

Cancer cell metastasis is the single most threatening occurrence of tumor progression and predicts patient prognosis as well as survival. Invasion can be regulated by the Met receptor tyrosine kinase (c-Met), integrin beta4, and the lipid enzyme, 12-Lipoxygenase (12-LOX). Therefore we sought to determine if beta4, c-MET and 12-LOX comprise a signaling axis. c-Met is implicated in cancer cell dissemination through regulation of invasion in EMT where cell-cell junctions are disturbed to allow motility. Furthermore, beta4 promotes cellular adhesion to the extracellular matrix through hemidesmosomes. However, the homeostatic signaling functions of beta4's cytoplasmic tail can be hijacked by growth factor …

Leptin Regulates Cd16 Expression On Human Monocytes In A Sex-Specific Manner, Joseph G. Cannon, Gyanendra Sharma, Gloria Sloan, Christiana Dimitropoulou, R. Randall Baker, Andrew Mazzoli, Barbara Kraj, Anthony Mulloy, Miriam Cortez-Cooper

School of Medical Diagnostics & Translational Sciences Publications

Fat mass is linked mechanistically to the cardiovascular system through leptin, a 16 kDa protein produced primarily by adipocytes. In addition to increasing blood pressure via hypothalamic-sympathetic pathways, leptin stimulates monocyte migration, cytokine secretion, and other functions that contribute to atherosclerotic plaque development. These functions are also characteristics of CD16-positive monocytes that have been implicated in the clinical progression of atherosclerosis. This investigation sought to determine if leptin promoted the development of such CD16-positive monocytes. Cells from 45 healthy men and women with age ranging from 20 to 59 years were analyzed. Circulating numbers of CD14++16++ monocytes, which are primary …

Molecular Phylogenetic Relationships Of North American Dermacentor Ticks Using Mitochondrial Gene Sequences, Kayla L. Perry

College of Graduate Studies: Theses & Dissertations

Dermacentor is a recently evolved genus of hard ticks (Family Ixodiae) that includes 36 known species worldwide. Despite the importance of Dermacentor species as vectors of human and animal disease, the systematics of the genus remain largely unresolved. This study focuses on phylogenetic relationships of the eight North American Nearctic Dermacentor species: D. albipictus, D. variabilis, D. occidentalis, D. halli, D. parumapertus, D. hunteri, and D. andersoni, and the recently re-established species D. kamshadalus, as well as two of the Neotropical Dermacentor species D. nitens and D. dissimilis (both formerly Anocentor). We sequenced portions of the mitochondrial …

Basic Features Of A Cell Electroporation Model: Illustrative Behavior For Two Very Different Pulses, Reuben S. Son, Kyle C. Smith, Thiruvallur R. Gowrishankar, P. Thomas Vernier, James C. Weaver

Bioelectrics Publications

Science increasingly involves complex modeling. Here we describe a model for cell electroporation in which membrane properties are dynamically modified by poration. Spatial scales range from cell membrane thickness (5 nm) to a typical mammalian cell radius (10 μm), and can be used with idealized and experimental pulse waveforms. The model consists of traditional passive components and additional active components representing nonequilibrium processes. Model responses include measurable quantities: transmembrane voltage, membrane electrical conductance, and solute transport rates and amounts for the representative "long" and "short" pulses. The long pulse-1.5 kV/cm, 100 μs-evolves two pore subpopulations with a valley at ~5 …

Cellular Regulation Of Extension And Retraction Of Pseudopod-Like Blebs Produced By Nanosecond Pulsed Electric Field, Mikhail A. Rassokhin, Andrei G. Pakhomov

Bioelectrics Publications

Recently we described a new phenomenon of anodotropic pseudopod-like blebbing in U937 cells exposed to nanosecond pulsed electric field (nsPEF). In Ca²⁺ -free buffer such exposure initiates formation of pseudopod-like blebs (PLBs), protrusive cylindrical cell extensions that are distinct from apoptotic and necrotic blebs. PLBs nucleate predominantly on anode-facing cell pole and extend toward anode during nsPEF exposure. Bleb extension depends on actin polymerization and availability of actin monomers. Inhibition of intracellular Ca²⁺ , cell contractility, and RhoA produced no effect on PLB initiation. Meanwhile, inhibition of WASP by wiskostatin causes dose-dependent suppression of PLB growth. Soon after …

Calcium-Mediated Pore Expansion And Cell Death Following Nanoelectroporation, Olga N. Pakhomova, Betsy Gregory, Iurii Semenov, Andrei G. Pakhomov

Bioelectrics Publications

Opening of long-lived pores in the cell membrane is the principal primary effect of intense, nanosecond pulsed electric field (nsPEF). Here we demonstrate that the evolution of pores, cell survival, the time and the mode of cell death (necrotic or apoptotic) are determined by the level of external Ca²⁺ after nsPEF. We also introduce a novel, minimally disruptive technique for nsEP exposure of adherent cells on indium tin oxide (ITO)-coated glass coverslips, which does not require cell detachment and enables fast exchanges of bath media. Increasing the Ca²⁺ level from the nominal 2–5 μM to 2 mM for …

Introduction To Fourth Special Issue On Electroporation-Based Technologies And Treatments, Damijan Miklavčič, Lluis M. Mir, P. Thomas Vernier

Bioelectrics Publications

This fourth special electroporation-based technologies and treatments issue of the Journal of Membrane Biology contains reports on recent developments in the field of electroporation by participants in the 7th International Workshop and Postgraduate Course on electroporation based technologies and treatments (EBTT 2013) held in Ljubljana, November 17–23, 2013. The 65 participants included faculty members, invited lecturers, special guests, and young scientists, and students from 16 countries. In addition to lectures on the fundamentals, this year’s sessions included talks on microbial inactivation by pulsed electric fields, modeling of intracellular electroporation, electroporation in food processing, and electrotransfer-facilitated DNA vaccination.

Molecular Mechanism Of Human Mismatch Repair Initiation, Sanghee Lee

Theses and Dissertations--Nutritional Sciences

DNA mismatch repair (MMR) is a highly conserved pathway that maintains genomic stability primarily by correcting mismatches generated during DNA replication. MMR deficiency leads to microsatellite instability (MSI), which is a hallmark of HNPCC (Hereditary Nonpolyposis Colorectal Cancer). Human mismatch repair is initiated by MutSα, a heterodimer of MSH2 and MSH6 subunits. Mismatch binding by MutSα triggers a series of downstream MMR events including interacting and communicating with other MMR proteins. The ATPase domain of MutSα is situated in the C-termini of its both subunits, and ATP binding is required for dissociation of MutSα from a mismatch. In eukaryotic cells, …

Regulation And Mechanistic Functions Of Caspase-9 Rna Splicing, Ngoc T. Vu

Theses and Dissertations

Caspase-9 has two splice variants, pro-apoptotic caspase-9a and anti-apoptotic caspase-9b, and dysregulation of caspase-9 splice variant ratio or expression of caspase-9b isoform has been linked to augmentation of the anchorage-independent growth and tumorigenic capacity of non-small cell lung cancer (NSCLC) cells. This study focuses on cell signaling pathway(s) regulating the alternative splicing of caspase-9 pre-mRNA and mechanistic roles of caspase-9b in a certain oncogenic/survival pathway. In regards to the former, we have identified hnRNP U as a novel splice-enhancer associated with exon 3 of caspase-9 (C9/E3). Moreover, hnRNP U binds specifically to C9/E3 at an RNA cis-element previously reported …

Structural And Functional Characterization Of The Mbd2-Nurd Co-Repressor Complex, Megha Desai

Theses and Dissertations

The MBD2-NuRD co-repressor complex is an epigenetic regulator of the developmental silencing of embryonic and fetal β-type globin genes in adult erythroid cells as well as aberrant methylation-dependent silencing of tumor suppressor genes in neoplastic diseases. Biochemical characterization of the MBD2-NuRD complex in chicken erythroid cells identified RbAp46/48, HDAC1/2, MTA1/2/3, p66α/β, Mi2α/β and MBD2 to comprise this multi-protein complex.

In the work presented in Chapter 2, we have pursued biophysical and molecular studies to describe a previously uncharacterized domain of human MBD2 (MBD2_IDR). Biophysical analyses show that MBD2_IDR is an intrinsically disordered region (IDR). Despite this inherent …