Biochemistry, Biophysics, and Structural Biology Commons^™

Investigating The Helicase Activity Of Methylated Vs Unmethylated Ded1, Hannah Lukow

Honors Theses

Ded1 is an RNA helicase protein of the DEAD-box subfamily in eukaryotic organisms (Sharma & Jankowsky, 2014) which can act as an activator or initiation factor, during translation (Hilliker et al., 2011). Ded1 has several functions in yeast including assembly of translational initiation factors, scanning the mRNA for the start codon, and unwinding any double stranded segments of mRNA with its helicase ability. Ded1 was discovered to be methylated at four arginine sites in vivo (Low et al., 2013), with a fifth methylation site being discovered recently (Low et al., 2020), however the purpose of such post-translational modifications is still …

Mechanism-Driven Approaches And Novel Constructs For High Purity Rna Synthesis, Kithmie Malagodapathiranage

Doctoral Dissertations

RNA is poised to revolutionize medicine. By simply changing the sequence, one therapeutic can be converted into a wholly new one, with little or no change in manufacturing and formulation. While a single mRNA vaccine produced at massive scale can treat billions, the re-codability of RNA will also enable the widespread growth of personalized medicines. T7 RNA polymerase is highly efficient at the synthesis of therapeutic RNA, but is known to produce unintended RNA impurities during synthesis. These products arise from the encoded RNA rebinding the enzyme such that its 3’ end serves as a primer for extension. This leads …

Analysis Of Ssa4 Reporter Expression By Q-Pcr, Susveen Sharanshi, Rebecca Adams

Belmont University Research Symposium (BURS)

The synthesis of genome-encoded proteins via mRNA translation is integral to cell survival. In eukaryotes, such as S. cerevisiae, the mRNA that is produced in the nucleus must be exported to the cytoplasm for translation to occur, and this process is highly regulated. Specifically, the export of mRNA occurs via travel through nuclear pore complexes (NPCs), which are selective doorways embedded in the nuclear envelope. During cellular stress, such as heat shock, the cell needs to regulate gene expression to permit survival, and mRNA export is one step at which this occurs. At these high temperatures, a cell’s proteins …

Role Of Rna Helicases In The Drosophila Germline, Patrick Blatt

Legacy Theses & Dissertations (2009 - 2024)

Gametogenesis, the process of creating egg or sperm, is required for launching successive generations of sexually reproducing organisms. The developmental milestones that occur during gamete production have been studied for decades and are of critical interest to gain insight to conserved features of human fertility. Drosophila has been used for over a century as an efficient research model and remains pivotal in uncovering fundamental biological paradigms. During Drosophila egg production, or oogenesis, several developmental transitions must be traversed to ensure completion of oogenesis including: Germline Stem Cell (GSC) maintenance and differentiation, mitotic and meiotic cell divisions, and production of maternally …

Alternative Polyadenylation Modulates Expression Of Pro-Fibrotic Proteins And Contributes To Lung Fibrosis, Junsuk Ko

Dissertations & Theses (Open Access)

Idiopathic pulmonary fibrosis (IPF) is a chronic lung disease which affects about 5 to 8 million individuals in the world. Despite the high prevalence, there is currently no cure for IPF, and the cause of this disease is still unclear. Our laboratory and collaborators have shown that nudix hydrolase 21 (NUDT21, which is also known as cleavage factor 25, CFIm25) is a key regulator of alternative polyadenylation (APA). NUDT21 depletion causes 3’UTR shortening via APA leading to enhanced mRNA stability and protein translation. This NUDT21 reduction promotes tumor growth in glioblastoma by enhancing expression of oncogenes. Cancer and IPF share …

Microrna Identification And Target Prediction In The Whitefly (Bemisia Tabaci), Alexis Aleman

Honors Theses

RNA interference, referred to as RNAi, is a biological phenomenon whereby knock-down of gene expression can be achieved through the use of RNA molecules, including small interfering RNAs (siRNAs) and microRNAs (miRNAs). The use of miRNAs is an endogenous pathway that results in the degradation of the miRNA strand’s complementary messenger RNA (mRNA), preventing the translation of the mRNA and therefore the production of the proteins necessary for gene expression. While RNAi is a biological phenomenon that occurs naturally, it is also a method that can be used and manipulated in the laboratory to try to control the expression of …

Parp1 Is A Versatile Factor In The Regulation Of Mrna Stability And Decay, Elena A. Matveeva, Lein F. Mathbout, Yvonne N. Fondufe-Mittendorf

Molecular and Cellular Biochemistry Faculty Publications

PARP1 is an abundant nuclear protein with many pleiotropic functions involved in epigenetic and transcriptional controls. Abundance of mRNA depends on the balance between synthesis and decay of a particular transcript. PARP1 binds RNA and its depletion results in increased expression of genes involved in nonsense-mediated decay, suggesting that PARP1 might be involved in mRNA stability. This is of interest considering RNA binding proteins play key roles in post-transcriptional processes in all eukaryotes. We tested the direct impact of PARP1 and PARylation on mRNA stability and decay. By measuring the half-lives of two PARP1-mRNA targets we found that the half-lives …

Human Rickettsial Pathogen Modulates Arthropod Organic Anion Transporting Polypeptide And Tryptophan Pathway For Its Survival In Ticks, Vikas Taank, Shovan Dutta, Amrita Dasgupta, Durland Fish, John F. Anderson, Hameeda Sultana, Girish Neelakanta

Biological Sciences Faculty Publications

The black-legged tick Ixodes scapularis transmits the human anaplasmosis agent, Anaplasma phagocytophilum. In this study, we show that A. phagocytophilum specifically up-regulates I. scapularis organic anion transporting polypeptide, isoatp4056 and kynurenine amino transferase (kat), a gene involved in the production of tryptophan metabolite xanthurenic acid (XA), for its survival in ticks. RNAi analysis revealed that knockdown of isoatp4056 expression had no effect on A. phagocytophilum acquisition from the murine host but affected the bacterial survival in tick cells. Knockdown of the expression of kat mRNA alone or in combination with isoatp4056 mRNA significantly affected A. phagocytophilum survival …

Hepatitis C Virus-Induced Monocyte Differentiation Into Polarized M2 Macrophages Promotes Stellate Cell Activation Via Tgf-Beta, Banishree Saha, Karen Kodys, Gyongyi Szabo

Gyongyi Szabo

BACKGROUND and AIMS: Monocyte and macrophage (MPhi) activation contributes to the pathogenesis of chronic hepatitis C virus (HCV) infection. Disease pathogenesis is regulated by both liver-resident MPhis and monocytes recruited as precursors of MPhis into the damaged liver. Monocytes differentiate into M1 (classic/proinflammatory) or M2 (alternative/anti-inflammatory) polarized MPhis in response to tissue microenvironment. We hypothesized that HCV-infected hepatoma cells (infected with Japanese fulminant hepatitis-1 [Huh7.5/JFH-1]) induce monocyte differentiation into polarized MPhis. METHODS: Healthy human monocytes were co-cultured with Huh7.5/JFH-1 cells or cell-free virus for 7 days and analyzed for MPhi markers and cytokine levels. A similar analysis was performed on …

Differential Activation Of Dead Box Rna Helicases Rhlb And Rhle By Hfq/Srnas And Their Target Mrnas, Amit Kumar

Wayne State University Theses

Number of small RNA (sRNA) gene regulators have mounted in E. coli over the years whereas the number of validated protein partners has not changed considerably. Hfq has remained the only well studied global regulatory partner of sRNAs in E. coli. However, direct or indirect involvement of other protein partners has always been speculated. Study from Blasi lab has shown that CsdA, one of the five DEAD-box RNA helicases of E. coli, is required for the DsrA mediated upregulation of rpoS under cold stress condition. Previous study from our lab has identified two other DEAD-box RNA helicases, RhlB and RhlE, …

Silica Nanoparticles For The Delivery Of Dna And Rnai In Cancer Treatment, Michael Aaron Vrolijk

Graduate College Dissertations and Theses

DNA and interfering RNA (RNAi) – short interfering RNA (siRNA) and micro RNA (miRNA) – are promising new cancer therapies, especially for drug resistant lines. However, they require a delivery system in vivo to prevent degradation and off target effects. Silica based nanoparticles, both solid and mesoporous, are a promising option due to their biocompatibility, ease of preparation and morphology control, reproducibility, and facile addition of functional groups including targeting ligands.

After a brief introduction to cancer treatment and review of the current nanoparticle treatments undergoing clinical trials, this thesis details the many methods explored over the past ten years …

Analysis Of Differential Mrna And Mirna Expression In An Alzheimer’S Disease Mouse Model, Amanda Hazy, Matthew Dalton

Other Undergraduate Scholarship

Research has shown that changes in gene expression play a critical role in the development of Alzheimer’s Disease (AD). Our project will evaluate genome-wide RNA expression patterns from brain and blood in an AD mouse model. This analysis will provide insight regarding the mechanisms of AD pathology as well as determine a possible diagnostic tool utilizing RNA expression patterns found in the blood as biomarkers for AD.

Translation Initiation Rate Determines The Impact Of Ribosome Stalling On Bacterial Protein Synthesis, Steven J. Hersch, Sara Elgamal, Assaf Katz, Michael Ibba, William Wiley Navarre

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Ribosome stalling during translation can be caused by a number of characterized mechanisms. However, the impact of elongation stalls on protein levels is variable, and the reasons for this are often unclear. To investigate this relationship, we examined the bacterial translation elongation factor P (EF-P), which plays a critical role in rescuing ribosomes stalled at specific amino acid sequences including polyproline motifs. In previous proteomic analyses of both Salmonella and Escherichia coli efp mutants, it was evident that not all proteins containing a polyproline motif were dependent on EF-P for efficient expression in vivo . The α- and β-subunits of …

Longitudinal Transcriptomic Dysregulation In The Peripheral Blood Of Transgenic Huntington’S Disease Monkeys, Jannet Kocerha, Yuhong Liu, David Willoughby, Kumaravel Chidamparam, Joseph Benito, Kate Nelson, Yan Xu, Tim Chi, Heidi Engelhardt, Sean Moran, Shang-Hsun Yang, Shi-Hua Li, Xiao-Jiang Li, Katherine Larkin, Adam Neumann, Heather Banta, Jinjing Yang, Anthony W. S. Chan

Jannet Kocerha

Background: Huntington’s Disease (HD) is a progressive neurodegenerative disorder caused by an expansion in the polyglutamine (polyQ) region of the Huntingtin (HTT) gene. The clinical features of HD are characterized by cognitive, psychological, and motor deficits. Molecular instability, a core component in neurological disease progression, can be comprehensively evaluated through longitudinal transcriptomic profiling. Development of animal models amenable to longitudinal examination enables distinct disease-associated mechanisms to be identified.

Results: Here we report the first longitudinal study of transgenic monkeys with genomic integration of various lengths of the human HTT gene and a range of polyQ repeats. With this unique group …

Section Abstracts: Biology With Microbiology And Molecular Biology

Virginia Journal of Science

Abstracts of the Biology with Microbiology and Molecular Biology Section for the 89th Annual Meeting of the Virginia Academy of Science, May 25-27, 2011, University of Richmond, Richmond VA.

The Role Of Upstream Sequences In Selecting The Reading Frame On Tmrna, Allen R. Buskirk, Mickey R. Miller, David W. Healey, Jonathan D. Dewey, Stephen G. Robison

Faculty Publications

tmRNA acts first as a tRNA and then as an mRNA to rescue stalled ribosomes in eubacteria. Two unanswered questions about tmRNA function remain: how does tmRNA, lacking an anticodon, bypass the decoding machinery and enter the ribosome? Secondly, how does the ribosome choose the proper codon to resume translation on tmRNA? According to the -1 triplet hypothesis, the answer to both questions lies in the unique properties of the three nucleotides upstream of the first tmRNA codon. These nucleotides assume an A-form conformation that mimics the codon-anticodon interaction, leading to recognition by the decoding center and choice of the …

In Vivo Role Of 20-Hydroxyecdysone In The Regulation Of The Vitellogenin Mrna And Egg Development In The American Dog Tick, Dermacentor Variabilis (Say), Deborah M. Thompson, Sayed M.S. Khalil, Laura A. Jeffers, Usha Ananthapadadmanaban, Daniel E. Sonenshine, Robert D. Mitchell, Christopher J. Osgood, Charles S. Apperson, R. Michael Roe

Biological Sciences Faculty Publications

Injection of the hormone 20-hydroxyecdysone (20-E) into partially fed (virgin) female adults of the American dog tick, Dermacentor variabilis, while they are attached and feeding on the rabbit host, initiated the expression of the vitellogenin (Vg) gene, and Vg protein secretion and uptake by the ovary. The induction of egg production by 20-E in this bioassay was dose dependent in the range of 1-50 times the concentration normally found in a replete, vitellogenic female. Ticks examined 4d after the 50x treatment were still attached to the host, had numerous enlarged vitellin-filled (brown) oocytes in their ovaries, but had not …