Medicine and Health Sciences Commons^™

Gamma Motor Neurons Express Distinct Genetic Markers At Birth And Require Muscle Spindle-Derived Gdnf For Postnatal Survival, Neil A. Shneider, Meghan N. Brown, Courtney A. Smith, James Pickel, Francisco J. Alvarez

Neuroscience, Cell Biology & Physiology Faculty Publications

Background: Gamma motor neurons (γ-MNs) selectively innervate muscle spindle intrafusal fibers and regulate their sensitivity to stretch. They constitute a distinct subpopulation that differs in morphology, physiology and connectivity from α-MNs, which innervate extrafusal muscle fibers and exert force. The mechanisms that control the differentiation of functionally distinct fusimotor neurons are unknown. Progress on this question has been limited by the absence of molecular markers to specifically distinguish and manipulate γ-MNs. Recently, it was reported that early embryonic γ-MN precursors are dependent on GDNF. Using this knowledge we characterized genetic strategies to label developing γ-MNs based on GDNF receptor expression, …

Structural Characterization Of The C-Terminus Of The Cochlear Motor Protein Prestin: Intrinsic Disorder Linked To Voltage Sensor Function, Michael S. Podgorski

Theses and Dissertations (ETD)

The sensitivity of mammalian hearing relies upon the ability to amplify sound. Outer hair cells (OHCs) in the cochlea have motile properties that support this function. The motor protein called prestin resides in the lateral wall of the OHCs. Prestin undergoes voltage-dependent conformational changes, which correlates to the movement of charge through the membrane or non-linear capacitance (NLC). This property underlies OHC electromotility. A mechanistic understanding of prestin’s function remains unknown. Previous research has tied prestin’s C-terminus (residues 499-799) to the voltage-dependent events that occur in the membrane. The folding and biochemical properties of prestin’s C-terminus were characterized. This information …

Identification Of The Downstream Effector Genes Involved In Hoxb4-Induced Expansion Of Hematopoietic Progenitor Cells, Jie Jiang

Theses and Dissertations (ETD)

Overexpression of HOXB4, a member of Homeobox transcription factor family, promotes expansion of hematopoietic stem and progenitor cells bothin vivo and in vitro. However, the molecular mechanisms underlying this effect are not well understood. In order to identify direct target genes of HOXB4 in primary murine hematopoietice progenitor cells, we induced HOXB4 function in lineage-negative, murine bone marrow cells, using a tamoxifen-inducibleHOXB4-ER^T2fusion protein. Seventy seven genes with differentially changed expression in early response to HOXB4 have been identified as candidate target genes. Among them, we show that Hemogen (Hemgn), encoding a nuclear protein specifically …

Expression And Function Of Hypoxia Inducible Factor-1 Alpha In Human Melanoma Under Non-Hypoxic Conditions, Caroline N. Mills, Sandeep S. Joshi, Richard M. Niles

Biochemistry and Microbiology

Background

Hypoxia inducible factor-1 alpha (HIF-1α) protein is rapidly degraded under normoxic conditions. When oxygen tensions fall HIF-1α protein stabilizes and transactivates genes involved in adaptation to hypoxic conditions. We have examined the normoxic expression of HIF-1α RNA and protein in normal human melanocytes and a series of human melanoma cell lines isolated from radial growth phase (RGP), vertical growth phase (VGP) and metastatic (MET) melanomas.

Results

HIF-1α mRNA and protein was increased in RGP vs melanocytes, VGP vs RGP and MET vs VGP melanoma cell lines. We also detected expression of a HIF-1α mRNA splice variant that lacks part …

Small-Molecule Screening Using A Human Primary Cell Model Of Hiv Latency Identifies Compounds That Reverse Latency Without Cellular Activation, Hung Chih Yang, Sifei Xing, Liang Shan, Karen O'Connell

University Faculty Publications and Creative Works

The development of highly active antiretroviral therapy (HAART) to treat individuals infected with HIV-1 has dramatically improved patient outcomes, but HAART still fails to cure the infection. The latent viral reservoir in resting CD4+ T cells is a major barrier to virus eradication. Elimination of this reservoir requires reactivation of the latent virus. However, strategies for reactivating HIV-1 through nonspecific T cell activation have clinically unacceptable toxicities. We describe here the development of what we believe to be a novel in vitro model of HIV-1 latency that we used to search for compounds that can reverse latency. Human primary CD4+ …

Ppar Agonists Down-Regulate The Expression Of Atp10c Mrna During Adipogenesis, A Peretich, Maria Cekanova Ms, Rndr, Phd, S Hurst, Sj Baek, Madhu Dahr

Faculty Publications and Other Works -- Biochemistry, Cellular and Molecular Biology

No abstract provided.

Ppar Agonists Down-Regulate The Expression Of Atp10c Mrna During Adipogenesis, A Peretich, Maria Cekanova Ms, Rndr, Phd, S Hurst, Sj Baek, Madhu Dahr

Maria Cekanova MS, RNDr, PhD

No abstract provided.

Accurate Molecular Classification Of Cancer Using Simple Rules., Xiaosheng Wang, Osamu Gotoh

Journal Articles: Genetics, Cell Biology & Anatomy

BACKGROUND: One intractable problem with using microarray data analysis for cancer classification is how to reduce the extremely high-dimensionality gene feature data to remove the effects of noise. Feature selection is often used to address this problem by selecting informative genes from among thousands or tens of thousands of genes. However, most of the existing methods of microarray-based cancer classification utilize too many genes to achieve accurate classification, which often hampers the interpretability of the models. For a better understanding of the classification results, it is desirable to develop simpler rule-based models with as few marker genes as possible.

METHODS: …

Foxp3-Expressing T Regulatory Cells (T-Regs) Increase With The Severity Of Active Disease In Chronic Hepatitis C, Adam D. Toll, Md, John L. Farber, Md

Department of Pathology, Anatomy, and Cell Biology Faculty Papers

The hepatitis C virus (HCV) leads to chronic disease in 80% of those infected and is associated with a chronic inflammatory response that is mediated by both cytokine producing (CD4+) and cytotoxic T cells (CD8+). FoxP3-expressing, CD4+, CD25+T cells (T-regs) are a subset of T lymphocytes that inhibit immune responsiveness and thereby control immunological reactions. Whether FoxP3+ T regulatory cell-mediated suppression is a factor in HCV persistence and/or the course of chronic liver injury has not been defined. In order to assess the association between these T regulatory cells and the severity of chronic hepatitis C, we evaluated liver biopsies …

The Production Of Antibody By Invading B Cells Is Required For The Clearance Of Rabies Virus From The Central Nervous System., D Craig Hooper, Timothy W Phares, Marzena J Fabis, Anirban Roy

Department of Cancer Biology Faculty Papers

BACKGROUND: The pathogenesis of rabies is associated with the inability to deliver immune effectors across the blood-brain barrier and to clear virulent rabies virus from CNS tissues. However, the mechanisms that facilitate immune effector entry into CNS tissues are induced by infection with attenuated rabies virus.

METHODOLOGY/PRINCIPAL FINDINGS: Infection of normal mice with attenuated rabies virus but not immunization with killed virus can promote the clearance of pathogenic rabies virus from the CNS. T cell activity in B cell-deficient mice can control the replication of attenuated virus in the CNS, but viral mRNA persists. Low levels of passively administered rabies …

Establishing The Purity Of Mononuclear Cell Preparations Using Morphology And Flow Cytometry, Adam Holtz, Bs, Adam D. Toll, Md, Susan W. Burkholder, Md, Alicia M. Carlin, Ba, Linda F. Blumstein, Bs, Kathleen A. Schroeder, Bs, Eric M. Behling, Md, Tina B. Edmonston, Md

Department of Pathology, Anatomy, and Cell Biology Faculty Papers

Context: Simple tandem repeat loci are used to track bone marrow engraftment using mononuclear buffy coat cells and T-cells. Poor isolation purity of these subpopulations can result in lower analytical sensitivity of the bone marrow engraftment assay by diluting the cell population in question with other nucleated cells. Validation of the mononuclear cell preparation can be performed by flow cytometry or by counting cell populations on the slide. Conclusions: Our results show that the purity of the Histopaque-1077 mononuclear cell preparation is excellent and that morphology may be sufficient to validate the mononuclear cell isolation method if flow cytometry is …

Intracellular Bacteria Encode Inhibitory Snare-Like Proteins., Fabienne Paumet, Jordan Wesolowski, Alejandro Garcia-Diaz, Cedric Delevoye, Nathalie Aulner, Howard A Shuman, Agathe Subtil, James E Rothman

Department of Microbiology and Immunology Faculty Papers

Pathogens use diverse molecular machines to penetrate host cells and manipulate intracellular vesicular trafficking. Viruses employ glycoproteins, functionally and structurally similar to the SNARE proteins, to induce eukaryotic membrane fusion. Intracellular pathogens, on the other hand, need to block fusion of their infectious phagosomes with various endocytic compartments to escape from the degradative pathway. The molecular details concerning the mechanisms underlying this process are lacking. Using both an in vitro liposome fusion assay and a cellular assay, we showed that SNARE-like bacterial proteins block membrane fusion in eukaryotic cells by directly inhibiting SNARE-mediated membrane fusion. More specifically, we showed that …

Role Of Mammalian Ecdysoneless In Cell Cycle Regulation., Jun Hyun Kim, Channabasavaiah B. Gurumurthy, Mayumi Naramura, Ying Zhang, Andrew T. Dudley, Lynn Doglio, Hamid Band, Vimla Band

Journal Articles: Genetics, Cell Biology & Anatomy

The Ecdysoneless (Ecd) protein is required for cell-autonomous roles in development and oogenesis in Drosophila, but the function of its evolutionarily conserved mammalian orthologs is not clear. To study the cellular function of Ecd in mammalian cells, we generated Ecd(lox/lox) mouse embryonic fibroblast cells from Ecd floxed mouse embryos. Cre-mediated deletion of Ecd in Ecd(lox/lox) mouse embryonic fibroblasts led to a proliferative block due to a delay in G(1)-S cell cycle progression; this defect was reversed by the introduction of human Ecd. Loss of Ecd led to marked down-regulation of E2F target gene expression. Furthermore, Ecd directly bound to Rb …

N-Glycosylation Status Of E-Cadherin Controls Cytoskeletal Dynamics Through The Organization Of Distinct Β-Catenin- And Γ-Catenin-Containing Ajs., Basem T Jamal, Mihai Nita-Lazar, Zhennan Gao, Bakr Amin, Janice Walker, Maria A Kukuruzinska

Department of Pathology, Anatomy, and Cell Biology Faculty Papers

N-glycosylation of E-cadherin has been shown to inhibit cell-cell adhesion. Specifically, our recent studies have provided evidence that the reduction of E-cadherin N-glycosylation promoted the recruitment of stabilizing components, vinculin and serine/threonine protein phosphatase 2A (PP2A), to adherens junctions (AJs) and enhanced the association of AJs with the actin cytoskeleton. Here, we examined the details of how N-glycosylation of E-cadherin affected the molecular organization of AJs and their cytoskeletal interactions. Using the hypoglycosylated E-cadherin variant, V13, we show that V13/β-catenin complexes preferentially interacted with PP2A and with the microtubule motor protein dynein. This correlated with dephosphorylation of the microtubule-associated protein …

Cd34 And Alpha-Smooth Muscle Actin Distinguish Cryptogenic Organizing Pneumonia From Secondary Bronchiolitis Obliterans-Organizing Pneumonia, Karen N. Wu, Md, Tajender S. Vasu, Md, John L. Farber, Md

Department of Pathology, Anatomy, and Cell Biology Faculty Papers

Secondary bronchiolitis obliterans-organizing pneumonia (BOOP) is a reaction to injury, forming granulation tissue with airspaces. Idiopathic cryptogenic organizing pneumonia (COP) and secondary BOOP are histologically similar. COP has been reported to show increased collagen in the granulation plugs and fewer myofibroblasts and capillaries. Using CD34 for capillaries and alpha-smooth muscle actin for myofibroblasts, we assessed these statins to differentiate COP from secondary BOOP.

The Elucidation Of Metabolic Pathways And Their Improvements Using Stable Optimization Of Large-Scale Kinetic Models Of Cellular Systems, Evgeni V. Nikolaev

Department of Pathology, Anatomy, and Cell Biology Faculty Papers

Metabolic engineering of cellular systems to maximize reaction fluxes or metabolite concentrations still presents a significant challenge by encountering unpredictable instabilities that can be caused by simultaneous or consecutive enhancements of many reaction steps. It can therefore be important to select carefully small subsets of key enzymes for their subsequent stable modification compatible with cell physiology. To address this important problem, we introduce a general mixed integer non-linear problem (MINLP) formulation to compute automatically which enzyme levels should be modulated and which enzyme regulatory structures should be altered to achieve the given optimization goal using non-linear kinetic models of relevant …

Detection Of Merkel Cell Carcinoma Polyomavirus In Mucosal Merkel Cell Carcinoma, Karen N. Wu, Md, Peter A. Mccue, Md, Zi-Xuan Wang, Phd, Agnieszka K. Witkiewicz, Md

Department of Pathology, Anatomy, and Cell Biology Faculty Papers

Case Report: A 61-year-old-woman with a past medical history significant for hypertension, bipolar disorder, obstructive sleep apnea and chronic obstructive pulmonary disease presented with diffusive lymphadenopathy suspicious for lymphoma. A lymph node in her right groin was biopsied by fine needle aspiration and showed atypical cells, but was not diagnostic. An excisional biopsy of the 2 x 2 cm lymph node demonstrated metastatic Merkel cell carcinoma (MCC). No skin lesions were detected. A CT scan revealed the presence of a large nasopharyngeal mass that following surgical removal was diagnosed as MCC. Treatment with etoposide and cisplatin and radiation were instituted. …

Pillows For Pain, Michael Lock

Michael Lock

No abstract provided.

Role Of Cd4+Cd25+ Regulatory T Lymphocytes In Experimental Toxoplasmosis, Sanjay Varikuti

Masters Theses & Specialist Projects

Toxoplasmosis is an important cause of congenital disease, and it is one of the most common opportunistic infections in patients with acquired immunodeficiency syndrome. The need for a reliable experimental model of this infection is crucial not only for achieving a better understanding of the patho-physiology of the disease, but also for developing better methods for evaluating new therapeutic regimens. The purpose of the present study was to investigate the role of CD4+CD25+ T regulatory lymphocytes in mice infected with Toxoplasma gondii. T regulatory (Treg) cells have been shown to play an important role in our immune system in …

Intra-Tumor Heterogeneity Of Mlh1 Promoter Methylation Revealed By Deep Single Molecule Bisulfite Sequencing., Katherine E Varley, David G Mutch, Tina B Edmonston, Paul J Goodfellow, Robi D Mitra

Department of Pathology, Anatomy, and Cell Biology Faculty Papers

A single tumor may contain cells with different somatic mutations. By characterizing this genetic heterogeneity within tumors, advances have been made in the prognosis, treatment and understanding of tumorigenesis. In contrast, the extent of epigenetic intra-tumor heterogeneity and how it influences tumor biology is under-explored. We have characterized epigenetic heterogeneity within individual tumors using next-generation sequencing. We used deep single molecule bisulfite sequencing and sample-specific DNA barcodes to determine the spectrum of MLH1 promoter methylation across an average of 1000 molecules in each of 33 individual samples in parallel, including endometrial cancer, matched blood and normal endometrium. This first glimpse, …

E-Science @ Umass: Anticipating And Supporting E-Science Activities At The University Of Massachusetts, Maxine G. Schmidt, Rebecca Reznik-Zellen

Maxine G Schmidt

In March of 2008 an Ad Hoccommittee of Science Librarians from the University of Massachusetts Five Campus System convened to discuss the challenges of e-science and prepare the Libraries for their role in e-science initiatives. Three primary outcomes intended to support e-science activities emerged from the work of the Ad Hoc committee.

Nerve Injection Of Viral Vectors Efficiently Transfers Transgenes Into Motor Neurons And Delivers Rnai Therapy Against Als., Rui Wu, Hongyan Wang, Xugang Xia, Hongxia Zhou, Chunyan Liu, Maria Castro, Zuoshang Xu

Department of Pathology, Anatomy, and Cell Biology Faculty Papers

RNA interference (RNAi) mediates sequence-specific gene silencing, which can be harnessed to silencing disease-causing genes for therapy. Particularly suitable diseases are those caused by dominant, gain-of-function type of gene mutations. In these diseases, the mutant gene generates a mutant protein or RNA product, which possesses toxic properties that harm cells. By silencing the mutant gene, the toxicity can be lessened because the amount of the toxic product is lowered in cells. In this report, we tested RNAi therapy in a mouse model for amyotrophic lateral sclerosis (ALS), which causes motor neuron degeneration, paralysis, and death. We used a transgenic model …

The Effect Of Endothelin-1 On The Expression Of Cdk Inhibitors P21 & P27 In Bovine Corneal Endothelial Cells, Lakshmi Reddy Bollu

Masters Theses & Specialist Projects

Mammalian corneal endothelial cells are considered to be non-proliferative due to the arrest of cells at the G1 phase of the cell cycle. The purpose of this study was to determine whether the down regulation of cyclin dependant kinase inhibitors (p21cip1 and p27kip1) levels by Endothelin-1 (ET-1), would overcome the G1 phase arrest and promote cell cycle progression and proliferation in cultured BCECs (Bovine corneal endothelial cells). BCECs were isolated from bovine corneas and cultured in DMEM supplemented with 10% serum. 5-Bromo 2-deoxy Uridine (BrdU) incorporation was determined in serum starved cultures in 24-well plates as a measure of cell …

Interaction With Lc8 Is Required For Pak1 Nuclear Import And Is Indispensable For Zebrafish Development., Christine M Lightcap, Gabor Kari, Luis E Arias-Romero, Jonathan Chernoff, Ulrich Rodeck, John C Williams

Department of Biochemistry and Molecular Biology Faculty Papers

Pak1 (p21 activated kinase 1) is a serine/threonine kinase implicated in regulation of cell motility and survival and in malignant transformation of mammary epithelial cells. In addition, the dynein light chain, LC8, has been described to cooperate with Pak1 in malignant transformation of breast cancer cells. Pak1 itself may aid breast cancer development by phosphorylating nuclear proteins, including estrogen receptor alpha. Recently, we showed that the LC8 binding site on Pak1 is adjacent to the nuclear localization sequence (NLS) required for Pak1 nuclear import. Here, we demonstrate that the LC8-Pak1 interaction is necessary for epidermal growth factor (EGF)-induced nuclear import …

Microarray-Based Cancer Prediction Using Soft Computing Approach., Xiaosheng Wang, Osamu Gotoh

Journal Articles: Genetics, Cell Biology & Anatomy

One of the difficulties in using gene expression profiles to predict cancer is how to effectively select a few informative genes to construct accurate prediction models from thousands or ten thousands of genes. We screen highly discriminative genes and gene pairs to create simple prediction models involved in single genes or gene pairs on the basis of soft computing approach and rough set theory. Accurate cancerous prediction is obtained when we apply the simple prediction models for four cancerous gene expression datasets: CNS tumor, colon tumor, lung cancer and DLBCL. Some genes closely correlated with the pathogenesis of specific or …

Decorin Is A Novel Antagonistic Ligand Of The Met Receptor., Silvia Goldoni, Ashley Humphries, Alexander Nyström, Sampurna Sattar, Rick T Owens, David J Mcquillan, Keith Ireton, Renato V Iozzo

Department of Pathology, Anatomy, and Cell Biology Faculty Papers

Decorin, a member of the small leucine-rich proteoglycan gene family, impedes tumor cell growth by down-regulating the epidermal growth factor receptor. Decorin has a complex binding repertoire, thus, we predicted that decorin would modulate the bioactivity of other tyrosine kinase receptors. We discovered that decorin binds directly and with high affinity (K(d) = approximately 1.5 nM) to Met, the receptor for hepatocyte growth factor (HGF). Binding of decorin to Met is efficiently displaced by HGF and less efficiently by internalin B, a bacterial Met ligand. Interaction of decorin with Met induces transient receptor activation, recruitment of the E3 ubiquitin ligase …

Regulation Of Pyruvate Dehydrogenase Kinase 4 By Thyroid Hormone / Role Of Peroxisome Proliferator Activated Receptor Gamma Coactivator-1 Alpha And Ccaat Enhancer Binding Protein, Ramy Naguib Attia

Theses and Dissertations (ETD)

Pyruvate dehydrogenase kinase 4 (PDK4) regulates pyruvate oxidation through the phosphorylation and inhibition of the pyruvate dehydrogenase complex (PDC). The PDC catalyzes the conversion of pyruvate to acetyl-CoA and it is an important control point in glucose and pyruvate metabolism. Previous studies had reported that PDK4 gene expression is induced by thyroid hormone (T3). These studies did not investigate the mechanisms by which T3 regulated PDK4 gene expression. I have examined the role of the thyroid hormone receptor (TR), transcriptional coactivators especially the peroxisome proliferator activated receptor gamma coactivator-1 (PGC-1α) and other transcription factors that act as accessory factors in …

Contribution Of Organic Cation Transporter 2 (Oct2) To Cisplatin-Induced Nephrotoxicity, Kelly K. Filipski

Theses and Dissertations (ETD)

Cisplatin is the most widely used anticancer agent; however, the cellular pharmacokinetics are poorly understood. Cisplatin is predominantly eliminated through the urine via active secretion and is associated with nephrotoxicity. Currently, prehydration therapy is employed to prevent toxic renal side effects; however it has not been completely ameliorated. The studies described herein aim to determine the mechanism in which cisplatin enters the kidney cell from the blood and how it is subsequently secreted into the urine. Organic cation transporter 2 (OCT2) and ABCC2 are highly expressed in the kidney on the basolateral and apical membrane, respectively. We determined the contribution …

Insights Into Btb-Cul3 Ubiquitin Ligases From The Structures Of Spop-Substrate Complexes, Min Zhuang

Theses and Dissertations (ETD)

Cullin-Ring ubiquitin ligases (CRLs) are E3 complexes that specifically recognize substrates through substrate adaptors. In the largest CRL subfamily, Cul3 binds a BTB domain, and a protein-interaction domain such as MATH recruits substrates for ubiquitination. Here we present biochemical and structural analyses of the MATH and BTB domain containing protein, SPOP, which regulates diverse signaling pathways. First, we identified a conserved SPOP Binding Consensus (SBC) motif in the transcriptional regulator Ci, the protein phosphatase Puc, and the chromatin component MacroH2A. The SBC motif specifically binds the MATH domain of SPOP, and is required for Puc ubiquitination in vitro and in …

Structural And Kinetics Studies Of The Enzyme Dihydropteroate Synthase And The Implications For Antibiotic Resistance, Katherine A. Ayers

Theses and Dissertations (ETD)

The goal of this study is to develop small molecule inhibitors of DHPS for the treatment of a variety of infectious diseases. According to the World Health Organization (WHO), infectious diseases kill more than 13 million people worldwide every year making it the second leading cause of death behind cardiovascular disease. The sulfonamide class of drugs has been in use since the 1930’s to treat many infectious agents and act by targeting the enzyme dihydropteroate synthase (DHPS) of the prokaryotic and lower eukaryotic folate pathway.

DHPS is an ideal drug target because humans do not synthesize folate de novo and …