Cell Biology Commons^™

Investigating The Escape Mechanism Of Sre Bearing Mrna Transcripts During Viral Host Shutoff, Daniel Macveigh-Fierro

Doctoral Dissertations

During viral infection, the virus and host clash for control over gene expression in an evolutionarily arms race that has raged for thousands of years. During lytic replication, Kaposi’s sarcoma associated herpesvirus (KSHV) triggers a massive RNA decay event known as host shut off. This causes over 70% of all RNA to be degraded suppressing the host antiviral response while freeing resources for viral replication. Our lab focuses on a subset of transcripts that escape from this viral degradation event using a cis acting 3’ UTR element known as a “SOX resistant element” or SRE. Although we have identified a …

Protein Trafficking In The Endoplasmic Reticulum Mediated By A Tpr-Containing Adapter Protein, Nathan P. Canniff

Doctoral Dissertations

The endoplasmic reticulum (ER) is a large, multifunctional organelle that acts as the entrance into the secretory pathway, which accounts for the folding and maturation of approximately one third of the human proteome. It is the largest organelle in most cell types and is comprised of a single lumen and a contiguous membrane. The ER is responsible for a multitude of roles including protein translocation, folding, maturation, quality control, and glycosylation to name a few. These processes are buoyed by a large collection of chaperones and cochaperones, the largest subfamily of which is characterized by the presence of tetratricopeptide repeat …

A Novel Transmembrane Ligand Inhibits T Cell Receptor Activation, Yujie Ye

Doctoral Dissertations

T lymphocytes (T cells) play essential roles in the adaptive immune system. Each mature T cell expresses one type of functional T cell receptor (TCR). The TCR recognizes antigens bound to the major histocompatibility complex (MHC) in antigen presenting cells. The resulting stimulation signal crosses the transmembrane domain of TCR and initiates downstream signaling cascades. The human immune system relies on TCRs to recognize a variety of pathogens. Normally, TCR can distinguish the self-antigens from pathogenic antigens. However, dysfunction or aberrant expression of TCRs causes different inflammatory and autoimmune diseases, which afflict millions of people annually (Chapter I). Current treatments …

Principles Of Aaa+ Proteases, Samar Mahmoud

Doctoral Dissertations

ATPases associated with diverse cellular activities (AAA+) proteases in bacteria help maintain protein homeostasis by degrading misfolded and regulatory proteins. While a handful of protein targets for these proteases have been identified in Caulobacter crescentus and other organisms, more research is needed to elucidate mechanisms that govern substrate specificity. In the second chapter of this thesis, I will elaborate on how AAA+ substrate specificity is less rigid than previous work has suggested and how limiting ATP or mutations can alter substrate preferences of the ClpXP protease. In the third chapter, I will highlight our efforts to use a quantitative proteomics …

Characterizing The Multifaceted Roles Of The Proteasomal Deubiquitinase Uch37 In Proteostasis, Heather A. Bisbee

Doctoral Dissertations

Cellular protein pools are maintained through the biological processes of synthesis, degradation and quality control. As the dysregulation of these processes has been implicated in diseases such as neurodegeneration and cancer, understanding their functions is critical for drug development. Modification of proteins with ubiquitin may direct them to the proteasome, a large cellular protease complex, for degradation. Yet, the proteasome contains three deubiquitinating enzymes (DUBs) which remove ubiquitin from proteins, potentially altering their fate. As each DUB recognizes specific ubiquitin linkages and architectures, their activity may regulate how the proteasome handles substrates in dynamic cellular contexts. In this work, we …

What I Talk About When I Talk About Integration Of Single-Cell Data, Yang Xu

Doctoral Dissertations

Over the past decade, single-cell technologies evolved from profiling hundreds of cells to millions of cells, and emerged from a single modality of data to cover multiple views at single-cell resolution, including genome, epigenome, transcriptome, and so on. With advance of these single-cell technologies, the booming of multimodal single-cell data creates a valuable resource for us to understand cellular heterogeneity and molecular mechanism at a comprehensive level. However, the large-scale multimodal single-cell data also presents a huge computational challenge for insightful integrative analysis. Here, I will lay out problems in data integration that single-cell research community is interested in and …

Tracking And Modulating Circadian Rhythms In Cell Culture Models, Sujeewa Sampath Lellupitiyage Don

Doctoral Dissertations

Circadian rhythms are 24–hour activity cycles that exist for all organisms, from yeast and bacteria to mammals. Epidemiological data has shown that disruptions to circadian rhythms are associated with various diseases, including cancers. Circadian rhythms are regulated at the cellular level by a "molecular clock,” whose oscillations in protein transcription and translation control a range of downstream pathways. However, the connections between altered rhythms and diseases at the molecular level is unclear. Therefore, in this project, I tracked circadian rhythms in a high-resolution manner and thoroughly analyzed them to more completely reflect circadian behaviors and clock functioning at the molecular …

Metabolism In The Colonocyte: Where Bacteria And Diet Meet, Bohye Park

Doctoral Dissertations

The host-microbiome interaction and the functions of microbial-derived metabolites, including butyrate, are important in colon health. Butyrate is the preferred energy source in colonocytes and has multiple physiological functions in the colon. A metabolic shift in colonocytes toward increased glucose utilization from butyrate oxidation is followed by several critical genetic modifications in cancerous colonocytes. Moreover, it has been suggested that the gut-microbiota composition is influenced by environmental factors, such as diets and the host's physiological status. Therefore, an understanding of the role of colonocyte metabolism toward impacting the host-microbiota commensal relationship would be an important step in understanding the functional …

Calmodulin Like 38 Is Required For Autophagy Of Hypoxia-Induced Cytoplasmic Rna Granules In Arabidopsis Thaliana, Sterling Field

Doctoral Dissertations

In response to the energy crisis resulting from submergence stress and hypoxia, the model plant Arabidopsis thaliana limits non-essential mRNA translation, and accumulates cytosolic stress granules. Stress granules are phase-separated mRNA-protein particles that partition transcripts for various fates: storage, degradation, or return to translation after stress alleviation. Another response by the plant cell to low oxygen stress is the induction of the turnover pathway autophagy. Stress granule regulation by autophagy occurs by a ‘granulophagy’ pathway in yeast and mammalian systems through which parts or whole stress granules are degraded. Whether this occurs in plants has not been investigated.

A connection …

Erecta Family Genes Regulate The Shoot Apical Meristem And Organ Formation, Daniel A. Degennaro

Doctoral Dissertations

Plants are sessile and must adjust their organ growth to their environments. A reservoir of stem cells in the shoot apical meristem (SAM) supplies cells for differentiation into organs. The SAM must balance organ production with stem cell maintenance. The ERECTA family (ERfs) encodes the leucine-rich repeat receptor-like kinases ERECTA (ER), ERECTA-LIKE 1 (ERL1), and ERL2. ERf signaling regulates organ initiation and stem cell maintenance. Results presented in this work include the following:

1) WUSCHEL (WUS) and CLAVATA3 (CLV3) make up a negative feedback loop to maintain SAM size. WUS and CLV3 expression localization is critical for …

The Histone Variant H2av Regulates Stress Responses And Tissue Development Through Interactions With Chromatin Insulator Proteins In Drosophila Melanogaster, James Ryan Simmons

Doctoral Dissertations

The ability of a cell to sense and respond to various forms of stress is essential to maintain integrity of the genome. Numerous pathways have been implicated in cellular responses to environmental and genotoxic stresses, often involving proteins and complexes that bind DNA directly to orchestrate changes in transcription and genome organization. Chromatin insulators describe a class of protein complex that bind specific sequences in the genome and work through two classically described functions: to restrict communication between enhancers and promoters through physical separation into different genomic domains and to prevent the spread of heterochromatin into euchromatic regions of the …

The Role Of Cdc42 Regulation In Cell Growth Pattern And Timing, Julie Robinson

Doctoral Dissertations

In fission yeast, polarized cell growth stops during division and resumes after cell separation at the end that existed in the previous generation. It is unclear how growth occurs specifically at this end after cell division. Via a pedigree analysis of mutant cells which display a unique polarity phenotype, we show the existence of a memory of growth. This memory of growth exists at the pre-existing cell end, which grew in the previous generation, and facilitates growth at that end in the next generation. This memory of growth gives the pre-existing end some advantage in a competition for active Cdc42, …

Biofabricated Constructs Of Carbon-Based Nanoparticles With Mesenchymal Stem Cells For Orthopedic Repair, Steven D. Newby

Doctoral Dissertations

Breakthroughs in tissue engineering are moving at a rapid rate especially in the regenerative bone biofabrication. Technology growth in the field of additive manufacturing (AM) such 3D bioprinting which provides the ability to create biocompatible 3D construct on which a cell source could be seeded is an encouraging substitute to autologous grafts.

This present research aims to biofabricate a construct for bone tissue engineering using AM technology. The biocompatible material was chosen corresponding to bones extracellular matrix (ECM) composition, which demonstrates an inorganic and organic development phase: Poly (lactic-glycolic acid) was chosen as the polymeric matrix of the compound, due …

The Role Of Med31 And Med12 In Directing Adipogenesis Of Human Adult-Derived Mesenchymal Stem Cells, Joseph Straub

Doctoral Dissertations

Selective gene expression is crucial in maintaining the self-renewing and multipotent properties of stem cells. Mediator is a large, evolutionarily conserved, multisubunit protein complex that modulates gene expression by relaying signals from cell type-specific transcription factors to RNA polymerase II. In humans, this complex consists of 30 subunits arranged in four modules: head, middle, tail, and kinase. In our introduction, we show the state of the field of Mediator study with a focus on the critical kinase module. In the following chapters, we used siRNA knockdowns to investigate the roles of the highly-conserved core subunit MED31 and the kinase module …

Substrate Selection In Endoplasmic Reticulum Protein Quality Control, Ben Adams

Doctoral Dissertations

Protein folding and maturation is a complex and error-prone process. Errors in this process may lead to deleterious effects ranging from non-functional single proteins to large-scale protein aggregation leading to cell death. It is essential for cellular function that protein misfolding does not occur unchecked, and therefore numerous chaperone systems exist within the cell. For the thousands of proteins which traffic through the secretory pathway, the primary site of folding and maturation is the endoplasmic reticulum (ER). Multiple chaperone pathways within the ER, generally termed ER protein quality control, must support the proper maturation process of these thousands of substrates. …

Engineering Stimuli-Responsive Polymeric Nanoassemblies: Rational Designs For Intracellular Delivery Of Biologics, Kingshuk Dutta

Doctoral Dissertations

Biologic drugs have gained enormous research attention in recent years as reflected by the development of multiple candidates to the clinical pipelines and an increased percentage of FDA approval. This is reasoned by the fact that biologics have been proven to deliver more predictive and promising benefits for many hard-to-cure diseases by ‘drugging the undruggable’ targets. However, the challenges associated with biologic drug development are multi-fold, viz, poor encapsulation efficacy, systemic instability, low cellular internalization and endosomal escape capability. Thus, it is essential to develop new molecular strategies that can not only address the associated drug delivery challenges, but also …

Micro-Physiological Models To Mimic Mucosal Barrier Complexity Of The Human Intestine In Vitro, Abhinav Sharma

Doctoral Dissertations

The mucosal barrier in the intestine is vital to maintain selective absorption of nutrients while protecting internal tissues and maintaining symbiotic relationship with luminal microbiota. This bio-barrier consists of a cellular epithelial barrier and an acellular mucus barrier. Secreted mucus regulates barrier function via in situ biochemical and biophysical interaction with luminal content that continually evolves during digestion and absorption. Increasing evidence suggests that a mucus barrier is indispensable to maintain homeostasis in the gastrointestinal tract. However, the importance of mucus barrier is largely underrated for in vitro mucosal tissue modeling. The major gap is the lack of experimental material …

Engineering Mesothelin-Binding Proteins As Targeted Cancer Diagnostics And Therapeutics, Allison Rita Sirois

Doctoral Dissertations

Cancer is a significant global health concern; and traditional therapies, including chemotherapeutics, are often simultaneously toxic yet ineffective. There is a critical need to develop targeted cancer therapeutics which specifically inhibit molecules or molecular pathways essential for tumor growth and maintenance. Furthermore, a targeted therapy is only effective when a patient's tumor expresses the molecular target; therefore, companion diagnostics, including molecular imaging agents, are a necessary counterpart of targeted therapies. Mesothelin (MSLN) is a cell surface protein overexpressed in numerous cancers, including triple-negative breast, pancreatic, ovarian, liver, and lung, with limited expression in normal tissues. Aberrant MSLN expression promotes tumor …

Iojap: Morphological And Physiological Phenotype Characterization In Arabidopsis, Thomas Jay Payne

Doctoral Dissertations

IOJAP protein is found in all organisms that contain a ribosome of bacterial origin. The majority of studies suggest that IOJAP plays a role in translation, although this has yet to be thoroughly investigated in plants. Using Arabidopsis thaliana, an extensive phenotype characterization of iojap mutants was performed. Many processes of plant growth were slightly impaired at optimal temperature (22˚C) but became severely hindered at low temperature (12˚C and 4˚C). These cold temperature defects manifested in an overall reduction of plant growth as well as variegation, chlorosis, leaf hyponasty, as well as reduced maximum quantum yield (F_v/F …

Involvement Of Calcium-Dependent Protein Kinases And Phosphatases In Sperm Capacitation-Associated Events, Bidur Paudel

Doctoral Dissertations

ABSTRACT To acquire fertilizing ability, mammalian sperm undergo a series of biochemical and physiological changes collectively known as capacitation^1,2. At the molecular level, capacitation is associated with a fast bicarbonate (HCO₃^-)-dependent activation of a unique type of soluble adenyl cyclase (sAC) and a consequent increase in cyclic AMP (cAMP) levels and PKA activation³. Activation of a cAMP/PKA pathway results in the phosphorylation of PKA substrates, which in turn initiates activation of several signaling cascades ultimately leading to an increase in phosphorylation on tyrosine residues (P-Tyr) of sperm axonemal proteins^4,5. Increase in …

Cation Channels, Ca2+ Homeostasis, And Fertility In The Mouse, Goli Ardestani

Doctoral Dissertations

In mammals, fully grown immature oocytes, also known as germinal vesicle (GV), are arrested at prophase of meiosis I. Upon increase of the luteinizing hormone (LH), GV oocytes resume meiosis and reach the metaphase II (MII) stage of the second meiosis, a process known as oocyte maturation. These mature oocytes, henceforth referred to as eggs, are ovulated arrested at the MII stage, from which they are released at the time of fertilization. As in many other cells, Ca²⁺ homeostasis plays a pivotal role in oocytes during maturation and following fertilization, and it is therefore carefully regulated in these cells. …

Tpr-Containing Proteins Control Protein Organization And Homeostasis For The Endoplasmic Reticulum, Jill Bradley-Graham

Doctoral Dissertations

The endoplasmic reticulum (ER) is a complex, multifunctional organelle comprised of a continuous membrane and lumen that is organized into several functional regions. It plays various roles including protein translocation, folding, quality control, secretion, calcium signaling, and lipid biogenesis. Cellular protein homeostasis is maintained by a complicated chaperone network, and the largest functional family within this network consists of proteins containing tetratricopeptide repeats (TPRs). TPRs are well-studied structural motifs that mediate intermolecular protein-protein interactions, supporting interactions with a wide range of ligands or substrates. Nine TPR-containing proteins have been shown to localize to the ER and control protein organization and …

De-Coding The Impact Of Evolved Changes In Gene Expression And Cellular Phenotype On Primate Evolution, Trisha Zintel

Doctoral Dissertations

The goal of the dissertation work outlined here was to investigate the influence of proximal processes contributing to evolutionary differences in phenotypes among primate species. There are numerous previous comparative analyses of gene expression between primate brain regions. However, primate brain tissue samples are relatively rare, and my results have contributed to the pre-existing data on more well-studied primates (i.e. humans, chimpanzees, macaques, marmosets) as well as produced information on more rarely-studied primates (i.e. patas monkey, siamang, spider monkey). Additionally, the primary visual cortex has not previously been as extensively studied at the level of gene expression as other brain …

Environmental Risk Factors For Inflammatory Bowel Disease: Triclosan And Other Consumer Antimicrobials, Katherine Z. Sanidad

Doctoral Dissertations

Inflammatory bowel disease (IBD) has become a serious health problem since the incidence and prevalence of IBD has dramatically increased throughout the world. There is evidence that environmental factors are primarily responsible for the increase of IBD, therefore, it is important to identify novel environmental risk factors to reduce the risk of IBD and its associated diseases. Antimicrobials used in consumer products might serve as environmental risk factors for IBD and its associated diseases. Triclosan (TCS), triclocarban (TCC), benzalkonium chloride (BAC), benzethonium chloride (BET), and chloroxylenol (PCMX) are widely used antimicrobial ingredients in consumer products and are ubiquitous contaminants in …

Spatiotemporal Modeling And Model Restructuration Approaches In Studies Of Intracellular Signalling Pathways, Md Shahinuzzaman

Doctoral Dissertations

"The main focus of the research is to understand the complex phenomena of cell transduction pathways and cell biology in a single cell. Mathematical modeling and experimental evaluation are widely used approaches for this kind of research. Firstly, A multiscale framework for protein-protein interaction has been established using Brownian dynamics algorithm. Sit specific feature, steric collision, diffusion, co-localization and complex formation with time and space has been included in this spatial modeling framework. By implementation of the time adaptive feature in this framework, the computation time reduces in an order of magnitude compared with traditional modeling framework. This multiscale Brownian …

Studies Of Dynein Anchoring Protein In Budding Yeast Saccharomyces Cerevisiae, Safia Omer

Doctoral Dissertations

Inheritance of equal genomic content is vital for cellular growth and survival. During cell division, the mitotic spindle is built to ensure equal chromosome segregation between dividing cells. Fidelity of segregation is achieved by the precise positioning of the mitotic spindle by force generators located at the cell cortex where they exert pulling on microtubule (MT). Cortical dynein, a minus end directed motor, generates pulling forces via microtubule end-on capture-shrinkage and lateral MT sliding mechanisms. In Saccharomyces cerevisiae, the dynein attachment molecule Num1 interacts with plasma membrane (PM), endoplasmic reticulum (ER), and mitochondria to facilitate spindle positioning across the …

Identifying Functional Components Of The Endoplasmic Reticulum Quality Control And Degradation Factor Edem1, Lydia Lamriben

Doctoral Dissertations

The ER Degradation-Enhancing Mannosidase-Like protein 1 (EDEM1) is a critical endoplasmic reticulum (ER) quality control factor involved in identifying and directing non-native proteins to the ER-associated protein degradation (ERAD) pathway. However, its recognition and binding properties have remained enigmatic since its discovery. Here we provide evidence for an additional redox-sensitive interaction between EDEM1 and Z/NHK that requires the presence of the single Cys on the α-1 antitrypsin ERAD clients. Moreover, this Cys-dependent interaction is necessary when the proteins are isolated under stringent detergent conditions, ones in which only strong covalent interactions can be sustained. This interaction is inherent to the …

The Interplay Between Polarity Regulators, Calcium, And The Actin Cytoskeleton During Tip Growth, Carlisle Bascom Jr

Doctoral Dissertations

Plant cell growth is a meticulously regulated process whereby the cell wall is selectively loosened to allow for turgor-pressure driven expansion. The rate of expansion must equal delivery of new material, or the cell will lyse. In many plant cells, this process happens diffusely around the cell. However, a number of plant cells have anisotropic shapes that require exquisite spatial control of secretion. One simple example of anisotropic patterning is tip growth; highly polarized cell expansion utilized by pollen tubes, root hairs, and moss protonemata. Investigating the role various molecules have in tip growth sheds light on how plant cells …

Characterization Of She1 Spindle Role Using Ceullular, Biochemical, And Biophysical Methods, Yili Zhu

Doctoral Dissertations

During development, metaphase spindles undergo large movement and/or rotation to determine the cell division axis. While it has been shown that spindle translocation is achieved by astral microtubules pulling and/or pushing the cortex, how metaphase spindle stability is maintained during translocation remains not fully understood. In budding yeast, our lab has previously proposed a model for spindle orientation wherein the mitotic spindle protein She1 promotes spindle translocation across the bud neck by polarizing cortical dynein pulling activity on the astral microtubules. Intriguingly, She1 exhibits dominant spindle localization throughout the cell cycle. However, whether She1 has any additional role on the …

Motor Protein Regulation In Mammalian Mitosis, Barbara Mann

Doctoral Dissertations

Developing and maintaining a multicellular organism relies on the fundamental biological process of cell division, which ensures that genetic material is equally segregated between daughter cells. During mitosis, cells completely rearrange their cytoskeleton into a bipolar spindle through the concerted efforts of microtubules, motor proteins, and microtubule-associated proteins, which cells must regulate spatially and temporally to prevent errors such as chromosomal missegregation: a major cause of cancer. Although the mitotic spindle is a validated target for chemotherapy drug resistance and redundant pathways have highlighted the need for new targets. It is therefore important to understand how proteins that help build …