Biochemistry, Biophysics, and Structural Biology Commons^™

Quantitative Analysis Of Endocytosis Of Macrophage Growth Factor Receptor By Molecular Imaging, Victor Georgiev, Emily Toering, Christopher Callahan

The Journal of Undergraduate Research

Endocytosis is a critical cellular process responsible for nutrient uptake, sampling the environment, and cellular signaling. Endocytosis of growth factor receptors is an important step in the regulation of receptor signaling functions. The macrophage growth factor receptor controls growth and differentiation of macrophages, however, the pathway by which the receptor is internalized is not well understood. The analysis showed that for the first time, knockout of endophilin A2 decreases the internalization of macrophage growth factor receptor in primary bone marrow macrophages. This shows that fast endophilin-mediated endocytosis plays a role in the control of the amount of growth factor receptors …

Diversity And Biocide Susceptibility Of Fungal Assemblages Dwelling In The Art Gallery Of Magura Cave, Bulgaria, Milena M. Mitova, Mihail Iliev, Alena Nováková, Anna A. Gorbushina, Veneta I. Groudeva, Pedro M. Martin-Sanchez

International Journal of Speleology

Magura Cave, north-western Bulgaria, possesses valuable rock-art paintings made with bat guano and dated from the period between the Eneolithic and Bronze Ages. Since 2008, the Art Gallery is closed to the general public in order to protect the paintings from vandalism, microclimatic changes caused by visitors and artificial illumination, and the consequent growth of fungi and phototrophs. Nevertheless, some tourist visits are allowed under the supervision of cave managers. This study provides the first scientific report on cultivable fungal assemblages dwelling different substrata in the Art Gallery. A total of 78 strains, belonging to 37 OTUs (Ascomycota 81%, Zygomycota …

Editing Of Misaminoacylated Trna Controls The Sensitivity Of Amino Acid Stress Responses In Saccharomyces Cerevisiae, Kyle Mohler, Rebecca Mann, Tammy J. Bullwinkle, Kyle W. Hopkins, Lin Hwang, Noah M. Reynolds, Brandon Gassaway, Hans-Rudolph Aerni, Jesse Rinehart, Michael Polymenis, Kym F. Faull, Michael Ibba

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Amino acid starvation activates the protein kinase Gcn2p, leading to changes in gene expression and translation. Gcn2p is activated by deacylated tRNA, which accumulates when tRNA aminoacylation is limited by lack of substrates or inhibition of synthesis. Pairing of amino acids and deacylated tRNAs is catalyzed by aminoacyl-tRNA synthetases, which use quality control pathways to maintain substrate specificity. Phenylalanyl-tRNA synthetase (PheRS) maintains specificity via an editing pathway that targets non-cognate Tyr-tRNA^Phe. While the primary role of aaRS editing is to prevent misaminoacylation, we demonstrate editing of misaminoacylated tRNA is also required for detection of amino acid starvation by …

A Novel Microrna-1207-3p/Fndc1/Fn1/Ar Regulatory Pathway In Prostate Cancer, Dibash K. Das, Olorunseun O. Ogunwobi

Publications and Research

Prostate cancer (PCa) is the second most common cause of cancer-specific deaths in the U.S. Unfortunately, the underlying molecular mechanisms for its development and progression remain unclear. Studies have established that microRNAs (miRNAs) are dysregulated in PCa. The intron-derived microRNA-1207-3p (miR-1207-3p) is encoded at the non-protein coding gene locus PVT1 on the 8q24 human chromosomal region, an established PCa susceptibility locus. However, miR-1207-3p in PCa had not previously been investigated. Therefore, we explored if miR-1207-3p plays any regulatory role in PCa. We discovered that miR-1207-3p is significantly underexpressed in PCa cell lines in comparison to normal prostate epithelial cells, and …

Utilizing Microfluidics Devices To Monitor Endothelial Cell Migration., Tsz Ming Jeremy Tsang, Jacob Lefler, Charles Voyton, Kenneth Christensen

Biomedical Engineering Western Regional Conference

Utilizing microfluidics devices to monitor endothelial cell migration.

Engineering Fret Biosensors For Microrna Presence/Absence Analysis, Nicholas E. Larkey, Sean M. Burrows

Biomedical Engineering Western Regional Conference

No abstract provided.

The Molecular Basis Of Talin2'S High Affinity Toward Β1-Integrin, Yaxia Yuan, Liqing Li, Yanyan Zhu, Lei Qi, Latifeh Azizi, Vesa P. Hytönen, Chang-Guo Zhan, Cai Huang

Molecular Modeling and Biopharmaceutical Center Faculty Publications

Talin interacts with β-integrin tails and actin to control integrin activation, thus regulating focal adhesion dynamics and cell migration. There are two talin genes, Tln1 and Tln2, which encode talin1 and talin2, and it is generally believed that talin2 functions redundantly with talin1. However, we show here that talin2 has a higher affinity to β1-integrin tails than talin1. Mutation of talin2 S339 to leucine, which can cause Fifth Finger Camptodactyly, a human genetic disease, completely disrupted its binding to β–integrin tails. Also, substitution of talin1 C336 with Ser enhanced the affinity of talin1, whereas substitution of talin2 S339 with …

Fibronectin And Androgen Receptor Expression Data In Prostate Cancer Obtained From A Rna-Sequencing Bioinformatics Analysis, Dibash K. Das, Thahmina Ali, Konstantinos Krampis, Olorunseun O. Ogunwobi

Publications and Research

Prostate cancer is the second most commonly diagnosed male cancer in the world. The molecular mechanisms underlying its development and progression are still unclear. Here we show analysis of a prostate cancer RNA-sequencing dataset that was originally generated by Ren et al. [3] from the prostate tumor and adjacent normal tissues of 14 patients. The data presented here was analyzed using our RNA-sequencing bioinformatics analysis pipeline implemented on the bioinformatics web platform, Galaxy. The relative expression of fibronectin (FN1) and the androgen receptor (AR) were calculated in fragments per kilobase of transcript per million mapped reads, and represented in FPKM …

Genome-Wide Dna Methylation Reprogramming In Response To Inorganic Arsenic Links Inhibition Of Ctcf Binding, Dnmt Expression And Cellular Transformation, Matthew Rea, Meredith Eckstein, Rebekah Eleazer, Caroline Smith, Yvonne N. Fondufe-Mittendorf

Molecular and Cellular Biochemistry Faculty Publications

Chronic low dose inorganic arsenic (iAs) exposure leads to changes in gene expression and epithelial-to-mesenchymal transformation. During this transformation, cells adopt a fibroblast-like phenotype accompanied by profound gene expression changes. While many mechanisms have been implicated in this transformation, studies that focus on the role of epigenetic alterations in this process are just emerging. DNA methylation controls gene expression in physiologic and pathologic states. Several studies show alterations in DNA methylation patterns in iAs-mediated pathogenesis, but these studies focused on single genes. We present a comprehensive genome-wide DNA methylation analysis using methyl-sequencing to measure changes between normal and iAs-transformed cells. …

Gestational Diabetes Mellitus And The Predisposing Factors, Syeda Sadia Fatima, Rehana Rehman, Faiza Alam, Sarosh Madhani, Bushra Chaudhry, Taseer Ahmed Khan

Department of Biological & Biomedical Sciences

Objective: To evaluate the occurrence of gestational diabetes mellitus and its association with demographic and anthropometric variables in pregnant women.
Methods: This cross-sectional study was conducted at the Aga Khan University Hospital, Abbasi Shaheed Hospital and Memon Hospital in Karachi, from February 2014 to December 2015, and comprised pregnant women who were screened by 75-g 2-hour oral glucose tolerance test, (24-28 weeks of gestation) and classified as per the criteria of the International Association of Diabetes and Pregnancy Study Group. Weight, body mass index and serum glycated haemoglobin levels were measured. Women with pre-gestational diabetes were excluded. SPSS 21 was …

Hmba Is A Putative Hsp70 Activator Stimulating Hexim1 Expression That Is Down-Regulated By Estrogen, Rati Lama, Chunfang Gan, Nethrie Idippily, Viharika Bobba, David Danielpour, Monica Montano, Bin Su Ph.D.

Chemistry Faculty Publications

Hexamethylene bis-acetamide inducible protein 1 (HEXIM1) is identified as a novel inhibitor of estrogen stimulated breast cell growth, and it suppresses estrogen receptor-a transcriptional activity. HEXIM1 protein level has been found to be downregulated by estrogens. Recently, HEXIM1 has been found to inhibit androgen receptor transcriptional activity as well. Researchers have used Hexamethylene bisacetamide (HMBA) for decades to stimulate HEXIM1 expression, which also inhibit estrogen stimulated breast cancer cell gene activation and androgen stimulated prostate cancer gene activation. However, the direct molecular targets of HMBA that modulate the induction of HEXIM1 expression in mammalian cells have not been identified. Based …

Biophysical Tools To Study Cellular Mechanotransduction, Ismeel Muhamed, Farhan Chowdhury, Venkat Maruthamuthu

Mechanical & Aerospace Engineering Faculty Publications

The cell membrane is the interface that volumetrically isolates cellular components from the cell's environment. Proteins embedded within and on the membrane have varied biological functions: reception of external biochemical signals, as membrane channels, amplification and regulation of chemical signals through secondary messenger molecules, controlled exocytosis, endocytosis, phagocytosis, organized recruitment and sequestration of cytosolic complex proteins, cell division processes, organization of the cytoskeleton and more. The membrane's bioelectrical role is enabled by the physiologically controlled release and accumulation of electrochemical potential modulating molecules across the membrane through specialized ion channels (e.g., Na^⁺, Ca²⁺, K^⁺ channels). …

Whole Genome Expression Profiling Associates Activation Of Unfolded Protein Response With Impaired Production And Release Of Epinephrine After Recurrent Hypoglycemia, J Kim, Edmund La Gamma, T Estabrook, N Kudrick, Bistra Nankova

NYMC Faculty Publications

Recurrent hypoglycemia can occur as a major complication of insulin replacement therapy, limiting the long-term health benefits of intense glycemic control in type 1 and advanced type 2 diabetic patients. It impairs the normal counter-regulatory hormonal and behavioral responses to glucose deprivation, a phenomenon known as hypoglycemia associated autonomic failure (HAAF). The molecular mechanisms leading to defective counter-regulation are not completely understood. We hypothesized that both neuronal (excessive cholinergic signaling between the splanchnic nerve fibers and the adrenal medulla) and humoral factors contribute to the impaired epinephrine production and release in HAAF. To gain further insight into the molecular mechanism(s) …

Therapeutic Exploitation Of Metabolic Checkpoints In Cancer Cells, Deven S. Patel

Dissertations, Theses, and Capstone Projects

During the G1 phase of the cell cycle, normal cells respond to growth factors and nutrients prior to entering S-phase to replicate its genome. We previously reported that the growth factor-mediated restriction point is distinguishable from a series of late G1 metabolic checkpoints mediated by essential amino acids (EAAs), the conditionally essential amino acid glutamine (Gln), and mTOR – the mammalian target of rapamycin. Mutations in genes encoding proteins that regulate G1 cell cycle progression are observed in virtually all cancers. We observed that cancer cells with K-Ras mutations bypass the late G1 Gln checkpoint when deprived of Gln and …

Chloride And Proton Binding In The E. Coli 2cl¯:1h+ Clc Exchanger, Catherine Chenal

Dissertations, Theses, and Capstone Projects

The CLC family of membrane proteins is a ubiquitously expressed class of proton and usually voltage-activated chloride transporters involved in a myriad of physiological functions. Crystallographic structures identify up to three chloride binding sites: external, central and intracellular located in the inner half of the trans-membrane domain. The CLC proteins, except for the kidney isoforms, are gated by the extracellular side-facing gating Glutamate (Ex, E148 in CLC-ec1, the E. coli exchanger), which is thought to undergo a conformational change upon protonation.

To sort out how the thermodynamic paths to H+ coupled Cl¯ binding and conformational change in CLC-ec1 at the …

Transcriptional And Post-Transcriptional Regulation Of Histone Variant H2a.Z During Sea Urchin Development, Mihai Hajdu

Dissertations, Theses, and Capstone Projects

Histone variant H2A.Z promotes chromatin accessibility at transcriptional regulatory elements and is developmentally regulated in metazoans. We characterize the transcriptional and post-transcriptional regulation of H2A.Z in the purple sea urchin Strongylocentrotus purpuratus. H2A.Z depletion by antisense translation-blocking morpholino oligonucleotides during early development causes developmental collapse, in agreement with its previously demonstrated general role in transcriptional multipotency. During H2A.Z peak expression in 24-h embryos, endogenous H2A.Z 3’ UTR sequences stabilize GFP mRNAs relative to those with SV40 3’ UTR sequences, although the 3’UTR of H2A.Z does not determine the spatial distribution of H2A.Z transcripts during embryonic and postembryonic development. We …

Lipid Sensing By Mammalian Target Of Rapamycin, Deepak Menon

Dissertations, Theses, and Capstone Projects

Mammalian target of Rapamycin (mTOR) is a protein kinase that integrates nutrient and growth factor signals to promote cellular growth and proliferation. mTOR exists in two complexes - mTORC1 and mTORC2 that are distinguished by their binding partners and signaling inputs. mTORC1 is responsive to growth factors, amino acids and glucose and is associated with Raptor; whereas, mTORC2 is responsive primarily to growth factors and is associated with Rictor. Raptor and Rictor confer substrate specificity to mTORC1 and mTORC2 respectively. Phosphatidic acid (PA), a lipid second messenger and a central metabolite for membrane phospholipid biosynthesis, is required for the stability …

An Assessment Of Potential False Positive E.Coli Pyroprints In The Cplop Database, Skyler A. Gordon

Master's Theses

The genetic information found in each species of organism is unique, and can be used as a tool to differentiate at the molecular level. This has caused rapid genotyping methods to become the cornerstone of a new area of research dependent on reading the genome as a form of identification. One of these specific identification methods, known as pyroprinting, relies on the small variation of DNA sequences within the same species to develop a unique, reproducible fingerprint. By simultaneously pyrosequencing multiple polymorphic loci within the ribosomal operons known as the intergenic transcribed spacers, a reproducible output is obtained, known as …

Multi-Instrument Comparison And Compilation Of Non-Methane Organic Gas Emissions From Biomass Burning And Implications For Smoke-Derived Secondary Organic Aerosol Precursors, Lindsay E. Hatch, Robert J. Yokelson, Chelsea E. Stockwell, Patrick R. Veres, Isobel J. Simpson, Donald R. Blake, John J. Orlando, Kelley C. Barsanti

Chemistry and Biochemistry Faculty Publications

Multiple trace-gas instruments were deployed during the fourth Fire Lab at Missoula Experiment (FLAME- 4), including the first application of proton-transfer-reaction time-of-flight mass spectrometry (PTR-TOFMS) and comprehensive two-dimensional gas chromatography-time-offlight mass spectrometry (GC×GC-TOFMS) for laboratory biomass burning (BB) measurements. Open-path Fourier transform infrared spectroscopy (OP-FTIR) was also deployed, as well as whole-air sampling (WAS) with onedimensional gas chromatography-mass spectrometry (GCMS) analysis. This combination of instruments provided an unprecedented level of detection and chemical speciation. The chemical composition and emission factors (EFs) determined by these four analytical techniques were compared for four representative fuels. The results demonstrate that the instruments are …

Quality Control By Isoleucyl-Trna Synthetase Of Bacillus Subtilis Is Required For Efficient Sporulation, Elizabeth Kermgard, Zhou Yang, Annika-Marisa Michel, Rachel Simari, Jacqueline Wong, Michael Ibba, Beth A. Lazazzera

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Isoleucyl-tRNA synthetase (IleRS) is an aminoacyl-tRNA synthetase whose essential function is to aminoacylate tRNA^Ile with isoleucine. Like some other aminoacyl-tRNA synthetases, IleRS can mischarge tRNA^Ile and correct this misacylation through a separate post-transfer editing function. To explore the biological significance of this editing function, we created a ileS(T233P) mutant of Bacillus subtilis that allows tRNA^Ile mischarging while retaining wild-type Ile-tRNA^Ile synthesis activity. As seen in other species defective for aminoacylation quality control, the growth rate of the ileS(T233P) strain was not significantly different from wild-type. When the ileS(T233P) strain was assessed for its ability to promote …

Evolution Of Complex Target Selex To Identify Aptamers Against Mammalian Cell-Surface Antigens, Prabodhika R. Mallikaratchy

Publications and Research

The demand has increased for sophisticated molecular tools with improved detection limits. Such molecules should be simple in structure, yet stable enough for clinical applications. Nucleic acid aptamers (NAAs) represent a class of molecules able to meet this demand. In particular, aptamers, a class of small nucleic acid ligands that are composed of single-stranded modified/unmodified RNA/DNA molecules, can be evolved from a complex library using Systematic Evolution of Ligands by EXponential enrichment (SELEX) against almost any molecule. Since its introduction in 1990, in stages, SELEX technology has itself undergone several modifications, improving selection and broadening the repertoire of targets. This …

Mechanisms Of Ikbke Activation In Cancer, Sridevi Challa

USF Tampa Graduate Theses and Dissertations

Cancer is the second leading cause of death in the USA and it is expected to surpass heart diseases making it important to understand the underlying mechanisms of cancer. The efforts to target single signaling molecule showed little success in increasing the patient survival and it can be due to increased compensation for cell survival by alternative pathway activations. Hence comprehensive understanding of the alternative signaling pathways may help us treat cancer better. Chronic inflammation is attributed to increased risk of cancer and emerging studies show the growing importance of both canonical and non-canonical IκB kinases such as IKKα, IKKβ, …

Development Of A Novel Method To Express And Purify Vaccinia Virus Early Transcription Factors A7 And D6 Using Bacteria, Omkar M. Gandbhir

Seton Hall University Dissertations and Theses (ETDs)

The eradication of Smallpox is one of the greatest human achievements. However other poxviruses still exist infecting humans (Molluscum Contagiosum, monkeypox, and Vaccinia) or livestock (cowpox, sheeppox). Though vaccines and anti-virals exist, the side effects are serious, and viral resistance has been detected. The genome amongst poxviruses is highly conserved, early gene promoters maintain a specific critical consensus region that can be targeted for development of novel broad spectrum therapeutics. Using Vaccinia early transcription factors (VETFs) a novel method of expressing and purifying VETF A7 and D6 was developed. A7 and D6 are required for transcription of early genes recruiting …

Effects Of Exposure To Quantum Dots In The External And Internal Environments Of Organisms, Kenton Layne Leigh

Student Theses and Dissertations

The use of nanotechnology in research and consumer products is increasing, but there are few studies regarding the safety of nanoparticles (NPs). NPs’ small size and increased surface area allow for unique physical and chemical properties not seen in bulk counterparts of the same material. Thus, NP interactions within environmental and biological matrices are not well understood. Specifically, quantum dots (QDs) have garnered increased attention due to their superior optical properties. However, little research on the effects of QDs on the environment is available. The studies described herein provide data regarding interactions of QDs in aquatic and terrestrial environments using …

Molecular Cloning And Characterization Of A Stilbenoid Prenyltransferase Gene Family In Peanut, Tianhong Yang

Student Theses and Dissertations

Peanut (Arachis hypogaea) produces prenylated stilbenoids as phytoalexins to fight against various pathogens. In addition, the prenylated stilbenoids exhibit several bioactivities with potential applications in human health. Despite their importance, the availability of prenylated stilbenoids is limited and their biosynthetic pathways remain to be elucidated. To date, stilbenoid prenyltransferase genes have not been identified from any plant species. Based on the prenyl unit position on the prenylated stilbenoids, it was hypothesized that different prenyltransferases are involved in the biosynthesis of distinct prenylated stilbenoids in peanut. To this end, an elicitor controlled stilbenoid bioproduction system was developed using hairy roots of …

Changing The Pathobiological Paradigm In Myelodysplastic Syndromes: The Nlrp3 Inflammasome Drives The Mds Phenotype, Ashley Basiorka

USF Tampa Graduate Theses and Dissertations

Note: Portions of this abstract have been previously published in the journal Blood, Basiorka et al. Blood. 2016 Oct 13, and has been reproduced in this manuscript with permission from the publisher.

Myelodysplastic syndromes (MDS) are genetically diverse hematopoietic stem cell malignancies that share a common phenotype of cytological dysplasia, ineffective hematopoiesis and aberrant myeloid lineage maturation. Apoptotic cell death potentiated by inflammatory cytokines has been considered a fundamental feature of MDS for over two decades. However, this non-inflammatory form of cell death cannot account for the inflammatory nature of these disorders. We report that a hallmark of lower-risk …

Ms-Read: Quantitative Measurement Of Amino Acid Incorporation, Kyle Mohler, Hans-Rudolph Aerni, Brandon Gassaway, Jiqiang Ling, Michael Ibba, Jesse Rinehart

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Ribosomal protein synthesis results in the genetically programmed incorporation of amino acids into a growing polypeptide chain. Faithful amino acid incorporation that accurately reflects the genetic code is critical to the structure and function of proteins as well as overall proteome integrity. Errors in protein synthesis are generally detrimental to cellular processes yet emerging evidence suggest that proteome diversity generated through mistranslation may be beneficial under certain conditions. Cumulative translational error rates have been determined at the organismal level, however codon specific error rates and the spectrum of misincorporation errors from system to system remain largely unexplored. In particular, until …

Engineering A Fluorescent Protease Sensor For In Vivo Protein Detection, Thomas C. Kinard

Honors Scholar Theses

This report details the results of an ongoing project to engineer a mutant form of Red Fluorescent Protein (RFP) variant mCherry that acts as a real-time in vivo protease sensor. The sought-after mutant only becomes fluorescent when exposed to Tobacco Etch Virus (TEV) Protease, this system’s model protease. This will be accomplished via the insertion of the TEV Protease Recognition Site (TEV-PRS) in such a position that, before cleavage, will prevent the protein from folding to fluorescent conformation, but upon cleavage, will allow for fluorescent conformation to occur. The cylindrical structure of the protein, composed of beta-pleated sheets, contains “loops” …

Deuterateer: A Tool For Quantifying Peptide Isotope Precision And Kinetic Proteomics, Bradley C. Naylor, Michael T. Porter, Elise Wilson, Adam Herring, Spencer Lofthouse, Austin Hannemann, Stephen R. Piccolo, Alan L. Rockwood, John C. Price

Faculty Publications

Motivation

Using mass spectrometry to measure the concentration and turnover of the individual proteins in a proteome, enables the calculation of individual synthesis and degradation rates for each protein. Software to analyze concentration is readily available, but software to analyze turnover is lacking. Data analysis workflows typically don’t access the full breadth of information about instrument precision and accuracy that is present in each peptide isotopic envelope measurement. This method utilizes both isotope distribution and changes in neutromer spacing, which benefits the analysis of both concentration and turnover.

Results

We have developed a data analysis tool, DeuteRater, to measure protein …

Novel Interconnections In Lipid Metabolism Revealed By Overexpression Of Sphingomyelin Synthase-1, Gergana M. Deevska, Patrick P. Dotson Ii, Alexander A. Karakashian, Giorgis Isaac, Mark Wrona, Samuel B. Kelly, Alfred H. Merrill Jr., Mariana N. Nikolova‑Karakashian

Physiology Faculty Publications

This study investigates the consequences of elevating sphingomyelin synthase 1 (SMS1) activity, which generates the main mammalian sphingolipid, sphingomyelin. HepG2 cells stably transfected with SMS1 (HepG2-SMS1) exhibit elevated enzyme activity in vitro and increased sphingomyelin content (mainly C22:0- and C24:0-sphingomyelin) but lower hexosylceramide (Hex-Cer) levels. HepG2-SMS1 cells have fewer triacylglycerols than controls but similar diacylglycerol acyltransferase activity, triacylglycerol secretion, and mitochondrial function. Treatment with 1 mm palmitate increases de novo ceramide synthesis in both cell lines to a similar degree, causing accumulation of C16:0-ceramide (and some C18:0-, C20:0-, and C22:0-ceramides) as well as C16:0- and C18:0-Hex-Cers. In these experiments, the …