Biochemistry, Biophysics, and Structural Biology Commons^™

The Role Of Secondary And Tertiary Structure In The Cap-Independent Translation Of Fgf-9 And Hif-1-Alpha, Amanda Michelle Whittaker

Dissertations, Theses, and Capstone Projects

Under normoxic conditions, eukaryotes initiate translation of RNA through eIF4E recognition of the 5’ cap. However, under cellular stress, eukaryotic translation must be initiated through a 4E-independent, or “cap-independent” mechanism, involving eukaryotic initiation factor 4G (eIF4G) binding directly to the 5’ untranslated regions (5’ UTR) of the RNA. eIF4G binding then recruits the ribosome to the transcript. While this mechanism is useful for translation of apoptotic transcripts and transcripts involved in cell survival, cap-independent translation is also utilized by oncogenic RNA for tumorigenesis. Previous work by our lab and others has categorized this recruitment and initiation mechanism as either internal-ribosome-entry-site …

Staying Sane In The Membrane: Neutral Sphingomyelinase 2 As A Master Regulator Of Plasma Membrane Ceramide, Zainuddin Quadri, Erhard Bieberich

Markey Cancer Center Faculty Publications

Ceramide, a key signaling sphingolipid in the plasma membrane, plays a pivotal role in fundamental cellular processes such as adhesion, polarity, and programmed cell death. The generation of plasma membrane ceramide is largely attributed to the activity of two types of sphingomyelinases: neutral sphingomyelinase 2 (nSMase2, Smpd3) and acid sphingomyelinase (aSMase, Smpd1). While many studies have explored ceramide generation following experimental activation of these enzymes, the mechanisms governing basal or steady- state ceramide levels have remained poorly understood. Using an innovative mass spectrometry approach developed in the Canals’ lab, the team has quantified the distinct contributions of nSMase2 and aSMase …

Tdp43 Autoregulation Gives Rise To Dominant Negative Isoforms That Are Tightly Controlled By Transcriptional And Post-Translational Mechanisms, Megan M. Dykstra, Kaitlin Weskamp, Nicolás B. Gómez, Jacob Waksmacki, Elizabeth Tank, M. Rebecca Glineburg, Allison Snyder, Emile Pinarbasi, Michael Bekier, Xingli Li, Morgan R. Miller, Jen Bai, Shameena Shahzad, Neha Nedumaran, Clare Wieland, Corey Stewart, Sydney Willey, Nikolas Grotewold, Jonathon Mcbride, John J. Moran, Aditya V. Suryakumar, Michael Lucas, Peter M. Tessier, Michael Ward, Peter K. Todd, Sami J. Barmada

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

The nuclear RNA-binding protein TDP43 is integrally involved in the pathogenesis of amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration (FTLD). Previous studies uncovered N-terminal TDP43 isoforms that are predominantly cytosolic in localization, prone to aggregation, and enriched in susceptible spinal motor neurons. In healthy cells, however, these shortened (s)TDP43 isoforms are difficult to detect in comparison to full-length (fl)TDP43, raising questions regarding their origin and selective regulation. Here, we show that sTDP43 is created as a by-product of TDP43 autoregulation and cleared by nonsense-mediated RNA decay (NMD). sTDP43-encoding transcripts that escape NMD are rapidly degraded post-translationally via the proteasome …

Investigating The Interactions Between Elm1 And Hsl7 In The Morphogenesis Checkpoint Of Saccharomyces Cerevisiae, Rachel Foster

Masters Theses & Specialist Projects

Cell shape is essential to understanding the normal regulatory processes that occur within the cell and can indicate there are abnormalities present when there is dysfunction in the cell's development, which can have broader implications regarding human disease. The morphogenesis pathway couples cell division to the timely formation of a daughter bud in Saccharomyces cerevisiae and consists of Elm1, Hsl1, Hsl7, and Swe1 proteins which interact with septins at the bud neck to provide stability for the bud and to facilitate cell division. Elm1 is needed for the recruitment of Hsl1 and subsequently Hsl7. Hsl7 will then tether Swe1 to …

Anti-Cancer Compounds Based On Natural Imidazole Compounds, Autumn Forsythe, Victoria Hardy, Kerri L. Shelton

CSU Tower Day

We are developing a comprehensive review on imidazole-based compounds of natural origin, the most common groups of compounds, and their properties, such as the imidazole ring. The intent of this presentation is to learn about the compound’s occurrence, general/predicted properties, and advantages. The review will be a fluid development comparing imidazole derivatives of natural origin with antinociceptive/lethal properties from the last 5 years. This review would include both purely extracted products but also those modified by synthesis.

Expression And Function Of Floral Symmetry Genes In The Regulation Of Flower Shape In Fedia Graciliflora, Theresa A. Mustacchio

Theses and Dissertations

Shifts between radial symmetry (actinomorphy) and bilateral symmetry (zygomorphy) in flowers have occurred multiple times independently within angiosperms (flowering plants). Evidence across angiosperms indicate the transcription factors RADIALIS (RAD), DIVARICATA (DIV), and CYCLOIDEA (CYC) play a major role in regulating floral symmetry and morphology. The CYC-like genes have been shown to shift their expression dorsally independently in nearly all shifts to bilaterally symmetrical flowers examined. Three core eudicot clades of CYC-like genes have been identified: CYC1, CYC2 and CYC3, with only CYC2 genes being shown to have a role in floral symmetry. We use the non-model plant, Fedia graciliflora (Caprifoliaceae) …

Cdh-3/Cadherin, Yap-1/Yap And Egl-44/Tead Promote Syx-2/Syntaxin And Eff-1 Fusogen-Mediated Phagosome Closure, Alec Whited, Aladin Elkhalil, Ginger Clark, Piya Ghose

Biology Theses - Archive

Physical interactions between cells, such as cell-cell junctions, can profoundly impact cell fate. A vital cell fate for normal development and homeostasis is programmed cell death. Cells fated to die must be efficiently cleared away via phagocytosis, and defects are associated with a variety of diseased states. Whether cell-cell physical associations affect programmed cell elimination has not been well-explored. Here we describe, in vivo, a cell-cell adhesion-driven signaling pathway that ensures compartment-specific cell clearance during development. We previously described the specialized cell death program “Compartmentalized Cell Elimination” (CCE) in the C. elegans embryo. During CCE, the tail-spike cell (TSC), a …

Characterizing And Targeting The Non-Catalytic Functions Of Phosphatase Of Regenerating Liver 3 (Prl-3) In Oncogenesis And Cancer Progression, Jeffery T. Jolly

Theses and Dissertations--Molecular and Cellular Biochemistry

Phosphatase of Regenerating Liver 3 (PRL-3) is frequently upregulated in various cancers and is associated with poor patient prognosis. Although traditionally studied for its phosphatase activity, PRL-3 also interacts with the CNNM family of magnesium transporters through its catalytic site, and these two functions are mutually exclusive at any given time. Most previous studies relied on a commonly used PRL-3 mutation that disrupts both phosphatase activity and CNNM binding, making it challenging to determine which function drives its oncogenic effects. To address this gap in the field, I utilized a panel of PRL-3 mutants that selectively disrupt either phosphatase activity …

Functional Analysis Of Ligand Binding Domain 2 Of Outer Surface Protein C Of Borreliella Burgdorferi, Gavin Z. Chambers

Theses and Dissertations

Lyme disease, caused by the spirochete Borreliella burgdorferi, is the most common vector-borne infection in North America. A critical determinant of early mammalian infection is outer surface protein C (OspC), a highly variable lipoprotein that is indispensable for spirochete survival but whose precise function remains unresolved. OspC has been proposed to mediate diverse roles, including complement evasion, adhesion, tissue dissemination, and antiphagocytic activity, supporting the view that it is a multifunctional virulence factor. Structural studies have identified two putative ligand-binding domains (LBD1 and LBD2), with LBD2 forming a solvent-exposed crown at the membrane-distal end of the OspC dimer.

To …

Systematic Review Of Immunohistochemistry Protocols, Arpitha Parthasarathy

Pitzer Senior Theses

Immunohistochemistry (IHC) is an essential technique in biological research that allows for the visualization and localization of proteins within tissue samples. This systematic review analyzes the most effective, efficient and reproducible protocols for IHC. It focuses on three critical steps: freezing, cryosectioning, and staining of skeletal muscle tissues in mice models. A total of 41, 44, and 48 articles on freezing, cryosectioning, and staining techniques, respectively, were analyzed, from reputable sources in biology like PubMed and Web of Science. Results showed the following. For freezing, the use of isopentane combined with liquid nitrogen appeared to be the best method, minimizing …

Characterization Of The Erv41-Erv46 Complex As A Retrograde Receptor For Misfolded Secretory Proteins, John Andrew Fuesler

Dartmouth College Ph.D Dissertations

The Endoplasmic Reticulum (ER), the site of secretory protein biosynthesis

provides a favorable environment to promote polypeptide folding, protein

oligomerization and export. Although the ER contains many chaperones and other

factors that assist in protein folding, this process is error prone. Eukaryotes have

evolved protein quality control checkpoints to maintain homeostasis in the secretory

pathway, however these processes are imperfect and may result in proteotoxicity

when toxic levels or aggregates of proteins arise. Mutations in proteins involved in

trafficking in the early secretory pathway are closely associated with disease states.

Therefore, it is of great interest to further understand how …

Bridging The Gap Between Structural And Cellular Biology: Insight Into The Organization Of Glycolytic Enzymes In Cells, Heather Hansen

Graduate Theses, Dissertations, and Problem Reports (ETD)

Glycolytic enzymes are spatially organized in the cytoplasm, and are shown to localize with cellular structures such as the plasma membrane, F-actin, mitochondria, or associate independently within the cytosol. The assembly of these enzymes is hypothesized to regulate pathway flux to meet the energetic needs of the cell. However, it is unclear whether all of these assemblies have the same mechanism of formation or function in the cell. We investigated the role of glycolytic enzyme assemblies in cells, using the glycolytic “gatekeeper” enzyme phosphofructokinase-1 (PFK1). PFK1 is extensively regulated by multiple mechanisms, including over 20 allosteric ligands, post translational modifications, …

Caspases Protect Against Hyperglycemia Induced Necroptosis-Like Cell Death In Brain Endothelial Cells, Aleena S. Mathew

Theses and Dissertations

Acute diabetes characterized by high spikes in blood glucose levels in association with severe hyperglycemia can lead to an increased risk of ischemic stroke in the brain. To better understand high blood glucose levels, hyperglycemia, in the brain we explored its effects on endothelial cells. Endothelial cells line the inner walls of blood vessels and are directly impacted in hyperglycemia. Brain endothelial cells are a major component of the blood brain barrier (BBB) which tightly regulates the brain’s microenvironment. The type of cell death such as apoptosis, caspase dependent, or necroptosis, caspase independent, can exacerbate diabetes associated complications. There is …

Characterization Of Dietary And Hormonal Influences On Renal Sodium Transporters And Channels In Rodent Models, Evelyn Chan

Scripps Senior Theses

Within kidneys, a diverse array of renal epithelial cells works to maintain electrolyte and water balance to regulate blood pressure and pH. These cells contain varying types of sodium transporters that adjust sodium reabsorption according to the body’s needs. Previous studies have observed sex-specific differences in renal sodium transporter regulation, which are thought to optimize reproduction in females. In this study, we performed high resolution microscopy and detailed characterization of cell type-specific expression and localization of major sodium transporters and channels in rodents in response to dietary and hormonal influences. Specifically, we examined the impact of dietary salt intake and …

Exploring The Effects Of Road Salt On Earthworm Species, Alea Mry Koston

Williams Honors College, Honors Research Projects

This study examines the effects of two road salts, sodium chloride (NaCl) and calcium magnesium acetate (CMA), on the health of two earthworm species: Eisenia fetida and Lumbricus terrestris. Over a 4-week period, approximately 168 earthworms were exposed to varying concentrations of either NaCl, CMA, or mixed ratios of both, with a control group included for comparison. The experiment involved monitoring the changes in earthworm weight and measuring their contraction rates to determine physiological responses. Statistical analysis revealed no significant differences in weight or activity between NaCl and CMA treatments for either species. However, higher concentrations of NaCl resulted in …

Biomechanics Of The Atherosclerotic Lipidome: (Un)Expected Statin Interactions And Membrane Remodelling, Ethan M. Fong

Pomona Senior Theses

Statins interact with lipid membranes in ways that shape their pharmacokinetics, off-target effects, and pleiotropic actions, yet these interactions' mechanical and biophysical consequences remain poorly understood. This thesis employs a combination of surface plasmon resonance (SPR), quartz crystal microbalance with dissipation monitoring (QCM-D), and rheological modelling to characterize how representative statins intercalate within biomimetic supported lipid bilayers (SLBs) of healthy and diseased composition. SPR reveals that lipophilic statins exhibit higher apparent affinities and slower dissociation kinetics, especially in cholesterol- and anionic lipid-rich membranes. QCM-D analyses demonstrate that statins alter membrane mass, dissipation, and interfacial stiffness in a statin- and membrane-dependent …

Temporal Dynamics Of Programmed Genome Rearrangement In Sea Lamprey Embryogenesis: A Deep Learning & Lightsheet Imaging Approach To Studying Early Developmental Transitions, Kasturi Rajandran

Theses and Dissertations--Biology

Programmed genome rearrangement (PGR) in the sea lamprey (Petromyzon marinus) leads to the elimination of ~0.5Gb (20%) of the germline genome during early embryogenesis, yet its timing relative to the maternal-to-zygotic transition (MZT) remains unclear. In this study, we imaged embryos cleared at 3-hour intervals (up to 72 hours post fertilization (hpf)) using high-resolution lightsheet fluorescence microscopy and a custom deep-learning image-analysis pipeline (~80% accuracy) to quantify nuclei and micronuclei counts. Micronuclei, which are the hallmarks of PGR first appear between 24 to 27hpf (vs ~36hpf previously reported) indicating PGR starts earlier than previously reported in the sea …

Developing Novel Therapeutic Strategies For Mcrc With A Brafv600e Mutation, Mariah Geisen

Theses and Dissertations--Toxicology and Cancer Biology

Colorectal cancer (CRC) is the second leading cause of cancer death worldwide. The overall incidence rates of CRC have decreased in those over 50, but there is an increasing incidence rate in those under 50. Risk factors for CRC include diet, family history, and a sedentary lifestyle. Early detection of CRC is key to patient survival and treatment response. Localized CRC has a survival rate of 90% and allows for surgical resection and chemotherapy. The 5-year survival for metastatic CRC (mCRC) is significantly reduced to around 14%. Despite early detection, 50% of patients will develop metastasis after surgical resection and …

Analysis Of Differential Gene Expression In Androgen-Independent Clones Derived From The Mycap Pca Cell Line, Jessie L. Tignor, Melanie Sinanian, Richard Inho Joh, David Gewirtz, Jason Reed

Undergraduate Research Posters

Androgen deprivation therapy (ADT) is a primary treatment strategy for prostate cancer (PCa), yet many tumors eventually develop androgen independence, leading to treatment resistance. To investigate the molecular changes underlying this transition, we analyzed differential gene expression in four androgen-independent (AI) clones derived from the Myc-CaP prostate cancer cell line using RNA sequencing. Gene expression profiles were compared to the parental Myc-CaP line, and differentially expressed genes (DEGs) were identified using DESeq2 and edgeR. The AI clones exhibited significant downregulation of senescence-associated genes, including Ezh2 and lamin B1, suggesting a loss of senescence-related chromatin repression. Additionally, upregulation of Wnt pathway …

Unleashing Serine Palmitoyltransferase In Test Tubes And Mice, Matthew W. Peart

Theses and Dissertations

Canonically known both for structural contributions to lipid bilayers and roles in cell signaling, the sphingolipids comprise a dynamic, multifaceted class of molecules which are studied to understand homeostatic cell biology as well as pathophysiology. Sphingolipids contribute to tissue-specific structures such as myelin, a multilayer membrane that allows for rapid conduction of action potentials along neuronal axons. All sphingolipids are downstream products of the rate-limiting and initiating enzyme in the de novo sphingolipid synthesis pathway, Serine palmitoyltransferase (SPT). Demonstrative of its status as the enzyme that catalyzes a crucial step in a biochemical pathway, SPT activity is strictly regulated. This …

Inductions Of Long-Term Potentiation With Biomimetic Electromagnetic Fields In Primary Neurons, Cooper E. Kansala

Theses and Dissertations (Comprehensive)

Endogenous brain rhythms guide neural communication and plasticity, raising the question of whether weak, temporally structured electromagnetic fields (EMFs) that mimic these rhythms can modulate neuronal activity in a targeted, biomimetic manner. This thesis investigates whether patterned EMFs designed to reflect the temporal structure of hippocampal long-term potentiation (LTP) are more effective at inducing neural plasticity than conventional sinusoidal fields or no stimulation at all. Cortical neurons cultured from postnatal rats were exposed to patterned EMFs for 30 minutes and assessed across three distinct experimental domains. In Chapter 2, microelectrode array (MEA) recordings revealed that the LTP-patterned field significantly increased …

Reversible Degradation Of Peptidoglycan By Lytic Transglycosylases, Aaron Devereaux

Theses and Dissertations (Comprehensive)

Antimicrobial resistance continues to be a burden on the global healthcare system with an estimated cost of billions of dollars and millions of deaths each year. Recently, there has been little to no development on new antibiotics to treat bacterial infection, and any that are developed become resisted to within a few years. Both Gram-positive and Gram-negative bacteria contain a mesh-like layer called peptidoglycan (PG) that surrounds their cells providing strength, cell shape, and protection from their environments. This layer is a polymer of N-acetylmuramic acid (MurNAc) and N-acetylglucosamine (GlcNAc) connected via a b-1,4-glycosidic bond, with each strand being cross-linked …

Regulation And Function Of Mec-2 Alternative Splicing In Single Cells, Canyon Calovich-Benne

Biological Sciences Theses and Dissertations

One mechanism for generating diversity at the single cell level is selective expression of a unique set of transcription factors (TFs) and RNA binding proteins (RBPs). These regulatory factors contribute to functions of cells by controlling cell fate, turning genes on and off, and processing of transcripts either co- or post-transcriptionally. Many genes, especially neuronal genes, have multiple different splicing events, promoters, and/or polyadenylation sites. Post-transcriptional coordination is in the infancy of being studied and the functional consequences of this coupling at the single cell level has yet to be explored. Here we show, mec-2 coordination of alternative transcription start …

P85Α Degradation Mediated By Ubiquitin E3 Ligase Fbxo21 Offers Therapeutic Potential In Leukemia, Kasidy K. Weber

Theses & Dissertations

Acute myeloid leukemia (AML) is a complex and heterogeneous disease characterized by the clonal expansion of myeloid blasts in the bone marrow. Despite significant therapeutic advances over the years, the prognosis for AML patients remains dismal, with high relapse rates and poor overall survival. The ubiquitin-proteasome system (UPS) plays a critical role in maintaining cellular homeostasis by regulating the degradation of proteins involved in essential processes such as cell cycle control, DNA repair, apoptosis, and various signaling pathways. Given this vital function, targeting ubiquitin E3 ligases within the UPS presents a promising strategy for developing more effective and targeted therapies …

Investigating The Expression Of C-Myc And Birc5 In Trail And Pbox-15 Treated Metastatic Prostate Cancer Cells, Corinne Kai Leng Chew, Hannah O'Toole, Daniela M. Zisterer, Seema M. Nathwani, Jade K. Pollock

SURE Journal: Science Undergraduate Research Experience Journal

Purpose Metastatic prostate cancer is associated with poor survival rate and limited treatment options. Overexpression of c-Myc and BIRC5 has been reported in prostate cancer cells which contribute to the development of the tumour environment. The specific tumour receptor mechanism of the death ligand TRAIL (tumour necrosis factor related apoptosis inducing ligand) has attracted remarkable therapeutic interest. However, many cancer cells are resistant to TRAIL monotherapy. A novel microtubule-targeting agent, PBOX-15 is known to exhibit anti-tumour activity in cancer cell lines including those with multi-drug resistance. Previous studies demonstrated that PBOX-15 is capable in sensitising cancer cells to TRAIL-mediated cell …

Comparing Multivalent Cross-Linked Sodium Alginate/ Montmorillonite Beads Containing Curcumin Or Cayenne Pepper As Drug Carriers And Their Antimicrobial Resistance, Shionainn Traynor, Shubham Sharma

SURE Journal: Science Undergraduate Research Experience Journal

The aim of the following work was to investigate and compare curcumin and cayenne pepper in microbeads composed of sodium alginate (SA) crosslinked with montmorillonite (MMT) to act as drug carriers and their antimicrobial resistance. The microbeads were prepared through ion-exchange in multivalent solutions of CaCl₂, MgCl₂ and AlCl₃. The microbeads were characterized by Scanning Electron Microscopy (SEM), Energy Dispersive X-ray analysis (EDX) and the time required for intercalation of curcumin and cayenne pepper with MMT. Simulated intestinal fluid (pH 7.3) and gastric fluid (pH 1.2) at 37°C were used for comparison in swelling studies. …

Editorial - Sure Journal Vol 6, Anne M. Friel, Eva Campion, Sinead Loughran

SURE Journal: Science Undergraduate Research Experience Journal

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Investigating Tumor Growth And Regulation In Supratentorial Ependymomas; The Impact Of Dlk1 And Egr1 Knockout, Om Sinojia

Honors Scholar Theses

Ependymomas (EPNs) are primary brain tumors that often arise from radial glial cells lining the ventricular system. Supratentorial ependymomas (ST-EPNs) are particularly aggressive, and understanding the molecular factors driving their growth is critical for developing targeted therapies. This study investigates the roles of DLK1 and EGR1, key regulators in cellular differentiation and tumorigenesis, in the development of ST-EPNs. We utilized genetically engineered mouse models to induce postnatal knockouts of DLK1 and EGR1 and evaluated tumor growth using histological and imaging techniques. Tumor area was quantified across multiple brain sections from both male and female mice. DLK1 knockout brains exhibited consistent …

Ecological And Pathological Applications Of The Heat Shock Response, Taylor Rae Stanley

Theses and Dissertations

The heat shock response (HSR) is a cytoprotective stress response pathway that regulates cellular proteostasis. The HSR is an evolutionarily conserved pathway that is essential for normal cellular functioning. Here, we explore the broad ecological and pathological impacts of the HSR. In an ecological context, we perform gene level analysis of the transcriptomes of two closely related sunfish. We found that the more invasive bluegill sunfish has gene expansions in two HSR gene families, the HSP70 family and the HSP90 family compared to the redear sunfish. These gene expansions were also observed in several other teleost fish species and were …

Apoa2 Increases Cholesterol Efflux Capacity To Plasma Hdl By Displacing The C-Terminus Of Resident Apoa1, Snigdha Sarkar, Jamie Morris, Youngki You, Hannah Sexmith, Scott E. Street, Stephanie M. Thibert, Isaac K. Attah, Chelsea M. Hutchinson Bunch, Irina V. Novikova, James E. Evans, Amy S. Shah, Scott M. Gordon, Jere P. Segrest, Karin E. Bornfeldt, Tomas Vaisar, Jay W. Heinecke, W. Sean Davidson, John T. Melchior

Saha Cardiovascular Research Center Faculty Publications

Abstract The ability of high-density lipoprotein (HDL) to promote cellular cholesterol efflux is a more robust predictor of cardiovascular disease protection than HDL-cholesterol levels in plasma. Previously, we found that lipidated HDL containing both apolipoprotein A-I (APOA1) and A-II (APOA2) promotes cholesterol efflux via the ATP-binding cassette transporter (ABCA1). In the current study, we directly added purified, lipid-free APOA2 to human plasma and found a dose-dependent increase in whole plasma cholesterol efflux capacity. APOA2 likewise increased the cholesterol efflux capacity of isolated HDL with the maximum effect occurring when equal masses of APOA1 and APOA2 coexisted on the particles. Follow-up …