EXCEED THE SPACE PROVIDED. Programmed cell death, or apoptosis, is a built-in, signal-induced process by which a cell self-destructs. It is a highly regulated mechanism that is critical for normal development, tissue homeostasis, and the elimination of pathogen- infected cells. In humans, misregulated programmed cell death is associated with tumorigenesis, neurodegenerative diseases, immunodeficiency, and viral pathogenesis. Although many evolutionarily conserved components of the cell death pathway have been identified, the molecular mechanisms involved in cellular regulation of apoptosis are still largely unknown. Since host cell apoptosis can limit virus multiplication, many viruses have evolved diverse strategies to regulate the cell death pathway. The proteins that mediate such viral intervention have provided key insight into the cell death program. The long term objective of this proposal is to define the molecular mechanisms by which apoptosis is regulated through the study of three baculovirus-encoded apoptotic regulators: P35, P49, and IAP. Our approach focuses on the use of baculovirus-infected insect cells as a powerful yet convenient system for molecular analysis of both the induction and suppression of apoptosis. Building on recent advances in the apoptosis field, we use integrated approaches in biochemistry, genetics, and cell biology to determine the molecular mechanism of P49 and IAP anti- apoptotic activity. We focus on P49's novel ability to inhibit an initiator caspase resistant to the pancaspase inhibitor P35 by defining the molecular determinants of caspase selectivity by both irreversible inhibitors. Utilizing the recently discovered capacity of baculoviruses to efficiently deliver apoptotic regulators to cultured Drosophila melanogaster cells, we identify the in vivo targets of P49 and P35 and define the caspase cascade in this model organism. We determine the molecular mechanism of virus IAP anti-apoptotic activity by characterizing the interactions between hybrid IAPs and cellular apoptotic effectors. In concert, we also investigate the functional significance of oligomerization for both viral and cellular IAPs by using novel dominant negative inhibitors. Collectively, these studies are expected to provide new and fundamental information on virus-host interactions and the regulation of programmed cell death in animals. Such knowledge will contribute to the development of therapeutic strategies for apoptosis-associated diseases. PERFORMANCE SITE ========================================Section End===========================================

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI040482-08
Application #
6835707
Study Section
Experimental Virology Study Section (EVR)
Program Officer
Johnson, David R
Project Start
1997-01-01
Project End
2007-12-31
Budget Start
2005-01-01
Budget End
2005-12-31
Support Year
8
Fiscal Year
2005
Total Cost
$251,986
Indirect Cost
Name
University of Wisconsin Madison
Department
Microbiology/Immun/Virology
Type
Other Domestic Higher Education
DUNS #
161202122
City
Madison
State
WI
Country
United States
Zip Code
53715
Byers, Nathaniel M; Vandergaast, Rianna L; Friesen, Paul D (2016) Baculovirus Inhibitor-of-Apoptosis Op-IAP3 Blocks Apoptosis by Interaction with and Stabilization of a Host Insect Cellular IAP. J Virol 90:533-44
Vandergaast, Rianna; Mitchell, Jonathan K; Byers, Nathaniel M et al. (2015) Insect inhibitor-of-apoptosis (IAP) proteins are negatively regulated by signal-induced N-terminal degrons absent within viral IAP proteins. J Virol 89:4481-93
Mitchell, Jonathan K; Byers, Nathaniel M; Friesen, Paul D (2013) Baculovirus F-box protein LEF-7 modifies the host DNA damage response to enhance virus multiplication. J Virol 87:12592-9
Taggart, David J; Mitchell, Jonathan K; Friesen, Paul D (2012) A conserved N-terminal domain mediates required DNA replication activities and phosphorylation of the transcriptional activator IE1 of Autographa californica multicapsid nucleopolyhedrovirus. J Virol 86:6575-85
Mitchell, Jonathan K; Friesen, Paul D (2012) Baculoviruses modulate a proapoptotic DNA damage response to promote virus multiplication. J Virol 86:13542-53
Vandergaast, Rianna; Schultz, Kimberly L W; Cerio, Rebecca J et al. (2011) Active depletion of host cell inhibitor-of-apoptosis proteins triggers apoptosis upon baculovirus DNA replication. J Virol 85:8348-58
Cerio, Rebecca J; Vandergaast, Rianna; Friesen, Paul D (2010) Host insect inhibitor-of-apoptosis SfIAP functionally replaces baculovirus IAP but is differentially regulated by Its N-terminal leader. J Virol 84:11448-60
Schultz, Kimberly L W; Friesen, Paul D (2009) Baculovirus DNA replication-specific expression factors trigger apoptosis and shutoff of host protein synthesis during infection. J Virol 83:11123-32
Schultz, Kimberly L W; Wetter, Justin A; Fiore, Diccon C et al. (2009) Transactivator IE1 is required for baculovirus early replication events that trigger apoptosis in permissive and nonpermissive cells. J Virol 83:262-72
Guy, Michael P; Friesen, Paul D (2008) Reactive-site cleavage residues confer target specificity to baculovirus P49, a dimeric member of the P35 family of caspase inhibitors. J Virol 82:7504-14

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