Biochemistry Commons^™

Effect Of Tamoxifen On Proteome Expression During In Vitro Myogenesis In Murine Skeletal Muscle C2C12 Cells, Emily A. Morris, Ahlenne Abreu, S. P. Scordilis

Biological Sciences: Faculty Publications

Tamoxifen (TMX), a selective estrogen receptor modulator, is commonly used in the treatment of hormone-responsive cancers. However, the effects of TMX in anabolic tissues harboring estrogen receptors, such as skeletal muscle, are poorly understood. We report a tandem mass-tag approach to TMX-treated myogenesis in C₂C₁₂ cells, a well-characterized model of in vitro murine skeletal muscle differentiation. A longitudinal analysis of >10,000 proteins identified in untreated C₂C₁₂ myogenesis revealed a novel subset of 1,062 myogenically regulated proteins. These proteins clustered into five distinct longitudinal expression trends which significantly overlap those obtained in similar analyses performed …

Identification And Characterization Of Silk Gland Specific Ugt34 Gene In Helicoverpa Zea, Courtney Nicole Wynn

Theses and Dissertations

Uridine diphosphate glycosyltransferase (UGT) is a multigene family of enzymes responsible for catalyzing glycosylation of small hydrophobic molecules. Recently, a genomic analysis of the corn earworm (Helicoverpa zea) identified 45 different UGT genes. We discovered a UGT gene (UGT34) showing high levels of expression exclusively in the silk gland tissue. The expression levels of UGT34 were analyzed in different developmental stages and silk gland sub-segments, revealing that UGT34 is generally expressed at all larval instar stages and largely expressed in the middle and posterior subsegments of the silk glands. The soybean looper (Chrysodeixis includens), another noctuid …

Upland Cotton And Nematodes: An Analysis Of Historical Resistance, Upcoming Threats, And Co-Inoculation Effects, Amanda Gaudin

Theses and Dissertations

Upland cotton (Gossypium hirsutum ) is an important fiber crop grown throughout the southern United States. Plant-pathogenic nematodes are worm-like animals that feed on the roots of most agronomic crops, including cotton. The southern root-knot nematode (Meloidogyne incognita, RKN) and the reniform nematode (Rotylenchulus reniformis, RN) cause significant yield losses in cotton every year. Current sources of resistance are effective but limited, therefore historical screenings of cotton accessions were revisited in search for novel resistance sources. None were identified but many of the screened accessions possessed markers of known root-knot nematode and reniform nematode resistance. …

Spatial Structure Formation By The Post-Transcriptional Gene Regulator Rsme In Pseudomonas Fluorescens Pf0-1, Anton Evans

Electronic Theses and Dissertations

Microorganisms are often found in microbial communities we call biofilms. Organisms living in these crowded environments have significant evolutionary pressure to retain access to the resources necessary to sustain life. My research uses the bacterium Pseudomonas fluorescens Pf0-1 to study how organisms evolve strategies to solve this crowding problem as aging colonies repeatedly generate mutant patches. These mutants expand the reach of the colony resulting in decreased local density as they push themselves up to the resource rich surface. These spatial structures result from social interactions between the mutant and the parental cells mediated through extracellular secretions, resulting in the …

Mechanisms Behind The Chaperone Activity Of Nucleic Acids, Theodore J. Litberg

Electronic Theses and Dissertations

Understanding the interplay between nucleic acids and protein aggregation is integral to the understanding of proteostasis, aging, and neurodegenerative disease progression. Nucleic acids are known to modulate the aggregation of PrP, tau, ⍺-synuclein, and other disease relevant proteins. Although the interactions between misfolded protein and nucleic acids can play a role in disease, this interaction may potentially be beneficial as well. Our group and others have shown nucleic acids can be powerful chaperones. Previous work has shown both RNA and DNA can prevent protein aggregation and RNA can pass off protein clients to the heat shock protein (Hsp) system. Here …

Reactive Oxygen Species And Weak Magnetic Fields: Unraveling The Mechanisms Of Planarian Regeneration And Tissue Growth, Luke James Kinsey

Dissertations

Reactive oxygen species (ROS) play a critical role in basic cellular behaviors such as apoptosis, proliferation, and differentiation. These molecules once thought to be harmful byproducts of metabolism, are also required for many complex and ubiquitous processes like wound healing, development, and regeneration. This study investigates the influence of ROS on planarian regeneration, with a particular focus on ROS dynamics in stem cell-mediated tissue growth. Data reveal that modulation of ROS levels leads to changes in cellular behaviors and tissue growth outcomes. Inhibition of ROS accumulation blocks stem cell proliferation and tissue growth, preventing planarian regeneration, while partial ROS accumulation …

Investigating The Effects Of E219 Mutations In M. Tuberculosis Indole-3-Glycerol Phosphate Synthase, Cinthya Moran

Theses, Dissertations and Culminating Projects

Tuberculosis (TB) is a serious infectious disease caused by Mycobacterium tuberculosis. Tuberculosis drug resistance is increasingly a problem, but recent research has provided results that suggest Indole-3-Glycerol Phosphate Synthase (IGPS) could be a potential drug target. IGPS is an enzyme that catalyzes the fourth step in the tryptophan biosynthesis pathway, a crucial process for optimal Mycobacterium tuberculosis growth. In this research, we studied IGPS from Mycobacterium tuberculosis (MtIGPS) to shed light on a particular conserved residue that has not been studied in depth in any IGPS homolog. My investigation focuses on understanding the role of the active site residue E219. …

Phenotypic And Transcriptomic Characterization Of Rice Snrk1 Mutants Developed By Crispr/Cas9 Mutagenesis, Maria Clara Faria Chaves

Graduate Theses and Dissertations

SnRK1 is a heterotrimeric protein kinase that is composed of a catalytic subunit (α) and two regulatory subunits (β and βγ), and it has a main role in regulating energy homeostasis in the plant by modulating anabolic and catabolic process. SnRK1 phosphorylates and alters the activities of enzymes involved in metabolism and regulates gene expression by altering the activity of chromatin-remodeling enzymes or the transcription factors. Rice contains three functional paralogs of SnRK1α: SnRK1αa (LOC_Os03g17980), SnRK1αb (LOC_Os08g37800), and SnRK1αc (LOC_Os05g45420). This study focused on the function of these SnRK1 paralogs by evaluating the phenotypic and transcriptomic characteristics and the disease …

Epitranscriptomic Regulation In Breast Cancer And Pcb-Induced Liver Disease., Belinda Petri

Electronic Theses and Dissertations

Post-transcriptional RNA modifications including N6-methyladenosine (m6A) regulate mRNA stability, splicing, and translation. My research examined m6A in two disease models: breast cancer (BCa) and non-alcoholic fatty liver disease (NAFLD). Acquired resistance to endocrine therapies (ET) develops in approximately 20% of BCa patients with estrogen receptor α positive (ER+) tumors following treatment. The mechanisms by which tumor cells evade ET are not completely understood. Using a cell line model, we investigated the role of an m6A reader protein, HNRNPA2B1 (A2B1) that is upregulated in ET-resistant ER+ BCa cells. Stable overexpression of A2B1 in ET-sensitive MCF-7 cells (MCF-7-A2B1), results in ET resistance, …

Determining The Existence And Regulation Of Microlipophagy In Primary Brown And White Adipocytes, Sara C. Arenas De Leon

Biomedical Sciences ETDs

Lipids are one of life’s four main macromolecules and provide essential functions to cells.

The degradation of lipid droplets and mobilization of lipids occurs through lipolysis. Emerging evidence demonstrates evidence of a selective form of autophagy in lipolysis. The process of microlipophagy has only recently been described, and many features of its regulation are still poorly understood. Our recent study showed that inhibiting mammalian target of rapamycin complex 1 (mTORC1) pathway by depletion of Raptor in adipocytes led to an influx of lysosomes and accumulation of lipid droplets within lysosome. Here, we expand on these previous findings. We were able …

X-Ray Crystal Structure Of E399q,E708q Ecm16 Double Mutant, Gileydis Guillama

Open Access Theses & Dissertations

The use of antibiotics has undoubtedly been a boon for humanity in combating infections and microbial threats. However, their widespread utilization has contributed to the emergence and spread of antibiotic resistance, which now poses a significant public health challenge. Streptomyces bacterium, produce diverse secondary metabolites with antibacterial, antifungal, antiviral, antitumoral, and immunosuppressant activities. Among these compounds is echinomycin, a nonribosomal peptide antibiotic synthesized by Streptomyces lasalocidi, which inhibits DNA replication and transcription by intercalating the DNA duplex at CpG steps. A gene called ecm16 was identified in the echinomycin biosynthetic gene cluster, which provides echinomycin self-resistance. Ecm16 recognizes DNA duplexes …

Structural Study Of Lsd13 And Lsd14 Type I Modular Polyketide Synthases From The Lasalocid A Biosynthesis Pathway, Dayan Viera

Open Access Theses & Dissertations

Bioactive natural products often possess a complex structure and have a large size which make their chemical synthesis highly challenging. To overcome this challenge, we are developing a biosynthetic method for production of natural products and their analogs. This technology is expected to decrease the synthesis time from years to weeks and reduce the cost of drug production by multiple orders of magnitude. We are investigating the biosynthesis pathway of lasalocid A, an anticancer antibiotic, to develop this technology. Specifically, we aim to characterize the enzymes structurally and biochemically in this pathway, thus paving the way for their rational engineering. …

Co-Targeting Nucleus Accumbens Associate 1 And Nf-Κb Signaling Synergistically Inhibits Melanoma Growth, Lixiang Gu, Xingcong Ren, Chrispus Ngule, Xiaofang Xiong, Jianxun Song, Zhiguo Li, Jin-Ming Yang

Markey Cancer Center Faculty Publications

Nucleus-accumbens-associated protein-1 (NAC1) is a cancer-related transcriptional factor encoded by the NACC1 gene, which is amplified and overexpressed in various human cancers and has been appreciated as one of the top potential cancer driver genes. NAC1 has therefore been explored as a potential therapeutic target for managing malignant tumors. Here, we show that NAC1 is a negative regulator of NF-κB signaling, and NAC1 depletion enhances the level of the nuclear NF-κB in human melanoma. Furthermore, the inhibition of NF-κB signaling significantly potentiates the antineoplastic activity of the NAC1 inhibition in both the cultured melanoma cells and xenograft tumors. This study …

Metabolic Disruption Induced By Mtor Signaling Pathway Inhibition In Regulatory T-Cell Expansion For Clinical Application, Roberto Gedaly, Gabriel Orozco, Alexandre P. Ancheta, Mackenzie Donoho, Siddharth Desai, Fanny Chapelin, Aman Khurana, Lillie J. Lewis, Cuiping Zhang, Francesc Marti

Markey Cancer Center Faculty Publications

Background: Regulatory T cell (Treg) therapy is considered an alternative approach to induce tolerance in transplantation. If successful, this therapy may have implications on immunosuppression minimization/withdrawal to reduce drug-induced toxicity in patients. The aim of this study was to assess the efficacy of the mTORC1/C2 inhibitor, AZD8055, in the manufacturing of clinically competent Treg cells and compare the effects with those induced by rapamycin (RAPA), another mTOR inhibitor commonly used in Treg expansion protocols.

Methods: Primary human Treg cells were isolated from leukapheresis product. Cell viability, expansion rates, suppressive function, autophagy, mitochondrial unfolded protein response (mitoUPR), and cell metabolic profile …

The S1p Receptor 1 Antagonist Ponesimod Reduces Tlr4-Induced Neuroinflammation And Increases Aβ Clearance In 5xfad Mice, Zhihui Zhu, Liping Zhang, Ahmed Elsherbini, Simone M. Crivelli, Priyanka Tripathi, Carmen Harper, Zainuddin Quadri, Stefka D. Spassieva, Erhard Bieberich

Markey Cancer Center Faculty Publications

Background Previously, we showed that the sphingosine-1-phosphate (S1P) transporter spinster 2 (Spns2) mediates activation of microglia in response to amyloid β peptide (Aβ). Here, we investigated if Ponesimod, a functional S1P receptor 1 (S1PR1) antagonist, prevents Aβ-induced activation of glial cells and Alzheimer’s disease (AD) pathology.

Methods We used primary cultures of glial cells and the 5XFAD mouse model to determine the effect of Aβ and Ponesimod on glial activation, Aβ phagocytosis, cytokine levels and pro-inflammatory signaling pathways, AD pathology, and cognitive performance.

Findings Aβ42 increased the levels of TLR4 and S1PR1, leading to their complex formation. Ponesimod prevented the …

Fbg Αc 389 – 402 Modulates Factor Xiii Crosslinking In The Fibrinogen Αc Region., Francis Dean Orlina Ablan

Electronic Theses and Dissertations

Fibrinogen (Fbg) is a coagulation protein critical for clot formation. Coagulation Factor XIII (FXIII) is a calcium-dependent transglutaminase that crosslinks reactive glutamines (Q) and lysines (K) between fibrin and other anti-fibrinolytic proteins. In the presence of Ca²⁺, FXIII could be activated non-proteolytically (FXIII-A°), or proteolytically by thrombin (FXIII-A*). Significant increases in clot stability and red blood cell retention are linked to FXIII activity in the fibrinogen αC region (Fbg Aα 221 – 610). This region contains several FXIII-reactive glutamines and lysines, as well as a binding site for FXIII-A* (Fbg αC 389 – 402) that includes a key …

Fatty Acids And Parasitism: Towards A Better Understanding Of Lipid Metabolism In Trypanosoma Brucei, Joshua Saliutama

All Dissertations

Trypanosoma brucei is an extracellular eukaryotic parasite that causes sleeping sickness in humans and cattle. As an extracellular parasite, T. brucei relies on the host’s nutrients to satisfy its growth requirements. The parasite is unusual because it does not uptake most of the host’s lipid species. Instead, T. brucei prefers to perform de novo synthesis of most lipid species. One of the lipid species that T. brucei can both uptake and synthesize is fatty acids. In my thesis work, I investigated the dynamics of fatty acid uptake, metabolism, and utilization of T. brucei. My work starts by determining the …

Biochemical Characterization Of Ssb1, Ssb2, Sycp3, And Meilb2 In Meiotic Recombination, Garrett Buzzard

All Dissertations

The genome is constantly at risk to damage from environmental and normal cellular processes. Double-strand breaks (DSBs) are one of the most deleterious forms of DNA damage as unrepaired DSBs ultimately result in cell death. Homologous recombination (HR) is a conserved DNA repair pathway that utilizes a homologous chromosome as a template for DNA synthesis to repair the DSBs relatively error-free. HR is active in both mitotic and meiotic cells but meiotic HR is initiated by the introduction of programmed DSBs into the genome. The process of HR repairs the DSBs and results in a physical connection between homologous chromosomes …

Methods To Study Activity Dependent Protein Synthesis In Autism Spectrum Disorder, Megan Webb, Karin F K Ejendal, Tamara L. Kinzer-Ursem

Discovery Undergraduate Interdisciplinary Research Internship

It is estimated by the World Health Organization that 1 in 100 children have autism spectrum disorder (ASD), a condition characterized by neurological differences that may impact a person’s learning or behavior. Clinically, ASD symptoms are alleviated with behavioral or pharmacological therapies, however, not all patients respond to these interventions. Deep brain stimulation (DBS) is a promising treatment of Parkinson’s disease that could also be effective in treating ASD. SynGAP1 is a protein involved in neuronal action that is crucial for regulating synaptic plasticity. Mutations in the SYNGAP1 gene causing haploinsufficiency can result in the manifestation of ASD symptoms. This …

Investigating Gene Looping In Yeast, Abbie Bangs, Justin Peters

Summer Undergraduate Research Program (SURP) Symposium

Saccharomyces cerevisiae, also known as brewer’s yeast, is the model organism we chose to interrogate a possible length dependence of gene looping. Gene looping occurs when the promoter and the terminator regions of a gene come together in a physical interaction to form a loop. RNA polymerase transcribes the gene at an amplified rate due to the gene loop (Figure 1), but little research exists as to what factors are important for this process, including the mechanics of the underlying DNA.

Analysis Of Biologically Active Secondary Metabolites From Fungi, Elizabeth E. Guevara, Kirk Manfredi

Summer Undergraduate Research Program (SURP) Symposium

Fungi produce secondary metabolites, which are organic compounds that allow the organism to be competitive in their environment. Secondary metabolites are not involved in growth and development, instead they affect ecological interactions to increase survivability. These compounds will often have antimicrobial properties, leading to their use in antibiotics. Isolating a compound with significant antimicrobial activity could lead to its use in therapeutic antibiotics. New antibiotics are needed to address the growing issue of antimicrobial resistant bacteria. This project explores the antimicrobial compounds produced by fungi collected Wind Cave National Park or carried by local insects. Accurate taxonomic identification of the …

Cobalt-Sulfur Coordination Chemistry Drives B12 Loading Onto Methionine Synthase, Romila Mascarenhas, Arkajit Guha, Zhu Li, Markus Ruetz, Sojin An, Javier Seravalli, Ruma Banerjee

Department of Biochemistry: Faculty Publications

Cobalt-sulfur (Co-S) coordination is labile to both oxidation and reduction chemistry and is rarely seen in Nature. Cobalamin (or vitamin B12) is an essential cobalt-containing organometallic cofactor in mammals, and is escorted via an intricate network of chaperones to a single cytoplasmic target, methionine synthase. In this study, we report that the human cobalamin trafficking protein, MMADHC, exploits the chemical lability of Co-S coordination, for cofactor off-loading onto methionine synthase. Cys-261 on MMADHC serves as the -axial ligand to cobalamin. Complex formation between MMADHC and methionine synthase is signaled by loss of the lower axial nitrogen ligand, leading to five-coordinate …

C···O And Si···O Tetrel Bonds: Substituent Effects And Transfer Of The Sif3 Group, Zhihao Niu, Qiaozhuo Wu, Qingzhong Li, Steve Scheiner

Chemistry and Biochemistry Faculty Publications

The tetrel bond (TB) between 1,2-benzisothiazol-3-one-2-TF₃-1,1-dioxide (T = C, Si) and the O atom of pyridine-1-oxide (PO) and its derivatives (PO-X, X = H, NO₂, CN, F, CH₃, OH, OCH₃, NH₂, and Li) is examined by quantum chemical means. The Si···O TB is quite strong, with interaction energies approaching a maximum of nearly 70 kcal/mol, while the C···O TB is an order of magnitude weaker, with interaction energies between 2.0 and 2.6 kcal/mol. An electron-withdrawing substituent on the Lewis base weakens this TB, while an electron-donating group has the opposite …

A Cost-Effective And Smart Sensing Tissue-Like Testbed For Surgical Training, Lysette Zaragoza

McNair Research Journal SJSU

A low-cost tissue-like testbed with six nodes of varying stiffness was developed for surgical training to provide pressure and force feedback data through image reception to human operators. Using SolidWorks, a 3D model of the box trainer housing was created. A pad for the distribution of smartsensing nodes and microcontroller connections was designed with open spaces for the respective components. The pad was 3D-printed with PLA filament. Flat piezoelectric pressure sensors were fabricated with conductive materials and velostat sensor material. Using static and dynamic analyses, three top sensors were chosen to be used in three pressure sensing nodes. A calibration …

Ranchsatdb: A Genome-Wide Simple Sequence Repeat (Ssr) Markers Database Of Livestock Species For Mutant Germplasm Characterization And Improving Farm Animal Health, Naveen Duhan, Simardeep Kaur, Rakesh Kaundal

Plants, Soils and Climate Student Research

Microsatellites, also known as simple sequence repeats (SSRs), are polymorphic loci that play an important role in genome research, animal breeding, and disease control. Ranch animals are important components of agricultural landscape. The ranch animal SSR database, ranchSATdb, is a web resource which contains 15,520,263 putative SSR markers. This database provides a comprehensive tool for performing end-to-end marker selection, from SSRs prediction to generating marker primers and their cross-species feasibility, visualization of the resulting markers, and finding similarities between the genomic repeat sequences all in one place without the need to switch between other resources. The user-friendly online interface …

Mechanisms Of Fibrillar Aggregation Of The Lens Small Heat Shock Proteins, Αa- And Αb-Crystallin, Russell James Mcfarland

Dissertations and Theses

The protein αB-crystallin represents the archetypical small heat-shock protein (sHSP), and together with αA-crystallin makes up the primary chaperone system in the eye lens, where they play a critical role in maintaining proteostasis. To achieve transparency, the lens undergoes a loss of organelles and both protein turnover halts, requiring that proteins remain functional for long periods of time as they accumulate chemical damage. The α-crystallins are highly abundant in the lens to stabilize proteostasis for multiple decades. Over time though, the small heat-shock proteins become overwhelmed by a high burden of destabilized clients and their own accumulated chemical damage. Mechanistic …

Pyrroline-5-Carboxylate Reductase-2 Promotes Colorectal Carcinogenesis By Modulating Microtubule-Associated Serine/Threonine Kinase-Like/Wnt/Β-Catenin Signaling, Raju Lama Tamang, Balawant Kumar, Sagar M. Patel, Ishwor Thapa, Alshomrani Ahmad, Vikas Kumar, Rizwan Ahmad, Donald F. Becker, Dundy Kiran Bastola, Punita Dhawan, Amar B. Singh

Department of Biochemistry: Faculty Publications

Background: Despite significant progress in clinical management, colorectal cancer (CRC) remains the third most common cause of cancer-related deaths. A positive association between PYCR2 (pyrroline-5-carboxylate reductase-2), a terminal enzyme of proline metabolism, and CRC aggressiveness was recently reported. However, how PYCR2 promotes colon carcinogenesis remains ill understood. Methods: A comprehensive analysis was performed using publicly available cancer databases and CRC patient cohorts. Proteomics and biochemical evaluations were performed along with genetic manipulations and in vivo tumor growth assays to gain a mechanistic understanding. Results: PYCR2 expression was significantly upregulated in CRC and associated with poor patient survival, specifically among PYCR …

Editorial: Rising Stars In Microbial Physiology And Metabolism: 2022, Nicole R. Buan, Ulrike Kappler

Department of Biochemistry: Faculty Publications

This Research Topic was initiated to highlight work by young authors, the rising stars in the field of microbial physiology and metabolism. Microbial physiology and metabolism is an interdisciplinary field of research that seeks to uncover how the metabolic pathways of a cell work together to determine cell fate and function, whether that be growth, replication, pathogenicity, predation, respiration and fermentation, homeostasis or death. Ultimately, researchers like the ones featured here seek to integrate biological information and physicochemical parameters to try to find the underlying rules governing microbial function so that we can understand, predict and design microbes and microbial …

Proteomic Approaches To Identify Unique And Shared Substrates Among Kinase Family Members, Charles Lincoln Howarth

Dartmouth College Ph.D Dissertations

Protein phosphorylation is a reversible post-translational modification that is a critical component of almost all signaling pathways. Kinases regulate substrate proteins through phosphorylation, and nearly all proteins are phosphorylated to some extent. Crucially, breakdown in phosphorylation signaling is an underlying factor in many diseases, including cancer. Understanding how phosphorylation signaling mediates cellular pathways is crucial for understanding cell biology and human disease.

Targeted protein degradation (TPD) is a strategy to rapidly deplete a protein of interest (POI) and is applicable to any gene that is amenable to CRISPR-Cas9 editing. One TPD approach is the auxin-inducible degron (AID) system, which relies …

Chloroquine And Cytosolic Galectins Affect Endosomal Escape Of Antisense Oligonucleotides After Stabilin-Mediated Endocytosis, Ekta Pandey, Ed Harris

Department of Biochemistry: Faculty Publications

Non-DNA-binding Stabilin-2/HARE receptors expressed on liver sinusoidal endothelial cells specifically bind to and internalize several classes of phosphorothioate antisense oligonucleotides (PS-ASOs). After Stabilin-mediated uptake, PS-ASOs are trafficked within endosomes (>97%–99%), ultimately resulting in destruction in the lysosome. The ASO entrapment in endosomes lowers therapeutic efficacy, thereby increasing the overall dose for patients. Here, we use confocal microscopy to characterize the intracellular route transverse by PS-ASOs after Stabilin receptor-mediated uptake in stable recombinant Stabilin-1 and -2 cell lines. We found that PS-ASOs as well as the Stabilin-2 receptor transverse the classic path: clathrincoated vesicle-early endosome-late endosome-lysosome. Chloroquine exposure facilitated endosomal …