Theses and Dissertations from UMD

Permanent URI for this communityhttp://hdl.handle.net/1903/2

New submissions to the thesis/dissertation collections are added automatically as they are received from the Graduate School. Currently, the Graduate School deposits all theses and dissertations from a given semester after the official graduation date. This means that there may be up to a 4 month delay in the appearance of a give thesis/dissertation in DRUM

More information is available at Theses and Dissertations at University of Maryland Libraries.

Browse

Filters

2004 - 200912010 - 20122

Settings

Subject: search.filters.subject.Cell Death

Search Results

Now showing 1 - 3 of 3
  • Thumbnail Image
    Item
    Protein permeability pathways in the mitochondrial outer membrane during apoptosis
    (2012) Vidyaramanan, Ganesan; Colombini, Marco; Cell Biology & Molecular Genetics; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)
    Apoptosis, a form of programmed cell death is a physiological, homeostatic process that guides the systematic removal of unwanted, dying or damaged cells from the body. A key step in apoptosis is the irreversible release of mitochondrial intermembrane space (IMS) proteins into the cytosol by a process called Mitochondrial Outer Membrane Permeabilization (MOMP). MOMP is regulated by a special class of proteins called Bcl-2 family proteins and a sphingolipid called ceramide. The pro-apoptotic Bcl-2 proteins, especially Bax and Bak can cooperate with ceramide to form channels in mitochondria that cause protein efflux during MOMP. The ability of ceramide to form protein-permeable channels in MOM is established. Bax and ceramide enhanced MOMP synergistically. The ability of Bax to stimulate ceramide channels was investigated. It was found that the apparent affinity of Bax for a ceramide channel increases with the ceramide channel size. The results indicate that Bax binds a small ceramide channel and drives its growth until the Bax molecule finds the best fit to the channel. This interaction between Bax and a ceramide channel does not require of the presence of other Bcl-2 proteins or mitochondrion-specific factors. The critical structural features of ceramide were investigated for their role in ceramide channel formation. Analogs of ceramide bearing modifications in the functional groups were analyzed for their ability to form channels to assess stability and also to interact with native ceramide to form channels to assess compatibility between interacting groups. The C1-hydroxyl was found to be indispensable for channel formation while the C3-hydroxyl was inconsequential to channel formation. The amide nitrogen with its ability to donate hydrogen was important for stability as methylating the nitrogen diminished the channel forming ability. Similarly, converting the carbonyl oxygen to a urea group, now more polar and a stronger hydrogen bond former resulted in more stable permeabilization. Changes to the hydrocarbon tails did not affect the ability to form channels. Phytoceramide, which has a C4 hydroxyl instead of the C4-C5 trans double bond formed stable channels but phytoceramide inhibited channel formation by ceramide suggesting incompatibility in structure. Bax activation involves translocation of Bax from the cytosol to the MOM, conformational changes and subsequent channel formation. All steps involved in Bax activation are not well-understood. We have used ionic strength as a modulating tool to dissect the different steps in Bax mediated MOMP. Increasing the ionic strength was found to delay formation of real-time permeability by Bax. Increasing the ionic strength resulted in smaller channels that grew in size slowly. The high permeability induced by low ionic strength was not reversed by raising the ionic strength suggesting that Bax channels are not in dynamic equilibrium with Bax monomers. Ionic strength also altered the sensitivity of Bax mediated MOMP to inhibition by Bcl-xL. Ionic strength, however did not affect Bax insertion into membranes. Thus, ionic strength presents a good diagnostic tool to modulate Bax mediated channel formation downstream of Bax insertion into membranes.
  • Thumbnail Image
    Item
    Role and Regulation of Autophagy During Developmental Cell Death in Drosophila Melanogaster
    (2010) McPhee, Christina Kary; Mount, Stephen M; Cell Biology & Molecular Genetics; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)
    Two prominent morphological forms of programmed cell death occur during development, apoptosis and autophagic cell death. Improper regulation of cell death can lead to a variety of diseases, including cancer. Autophagy is required for survival in response to starvation, but has also been associated with cell death. It is unclear how autophagy is regulated under specific cell contexts in multi-cellular organisms, and what may distinguish autophagy function during cell survival versus cell death. Autophagic cell death is characterized by cells that die in synchrony, with autophagic vacuoles in the cytoplasm, and phagocytosis of the dying cells is not observed. However, little is known about this form of cell death. Autophagic cell death is observed during mammalian development, during regression of the corpus luteum and involution of the mammary and prostate glands. Autophagic cell death is also observed during development of the fruitfly Drosophila melanogaster, during larval salivary gland cell death. Drosophila is an excellent genetic model system to study developmental cell death in vivo. Cells use two main catabolic processes to degrade and recycle cellular contents, the ubiquitin/proteasome system (UPS) and autophagy. Here I investigate the role of the UPS and autophagy in developmental cell death using Drosophila larval salivary glands as an in vivo model. Proteasome inhibitors are being used in anti-cancer therapies; however the cellular effects of proteasome inhibition have not been studied in vivo. Here I demonstrate that the UPS is impaired during developmental cell death in vivo. Taking a proteomics approach to identify proteins enriched in salivary glands during developmental cell death and in response to proteasome impairment, I identify several novel genes required for salivary gland cell death, including Cop9 signalsome subunit 6 and the engulfment receptor Draper. Here I show that the engulfment receptor Draper is required for salivary gland degradation. This is the first example of an engulfment factor that is autonomously required for self-clearance. Surprisingly, I find that Draper is cell-autonomously required for autophagy during cell death, but not for starvation-induced autophagy. Draper is the first factor to be identified that genetically distinguishes autophagy that is associated with cell death from cell survival.
  • Thumbnail Image
    Item
    Identification of autophagic cell death and implications for therapy
    (2004-05-04) Alva, Ajjai Shivaram; Baehrecke, Eric H; Cell Biology & Molecular Genetics
    Autophagy is an evolutionarily conserved mechanism of bulk protein and organelle degradation that requires the ATG class of genes. Although autophagy has been frequently observed in dying cells in several species, a causative role for autophagy in cell death has not been demonstrated. We show that inhibition of caspase-8 in mouse L929 fibroblast cells causes cell death with the morphology of autophagy. Autophagic cell death in L929 cells is dependent on ATG genes and involves the receptor interacting protein (RIP) and the activation of the MAP kinase kinase 7(MKK7) - Jun N-terminal kinase (JNK) - cJUN pathway. We also show that autophagy occurs in many primary human tumors including cancer of the breast, lung and pancreas. Our findings validate autophagic cell death and might explain the role of autophagy in development, viral infections, neurodegenerative diseases and cancer.