Dorsoventral polarity of the Drosophila embryo is established by local activation of the Toll signaling pathway. The membrane receptor Toll appears to be activated by an extracellular ligand that is localized to the ventral side of the embryo. The Toll ligand is generated by a proteolytic processing reaction which requires four members of the serine protease family encoded by the nudel, gastrulation defective (gd), snake, and easter genes. Earlier genetic studies suggested that these proteases act in a sequential activation cascade in which the final protease processes the Toll ligand precursor, and that activation of proteases in this cascade is ventrally localized, thereby spatially restricting production of the Toll ligand. Our more recent studies suggest that the pathway producing the Toll ligand is regulated at multiple steps so that the Toll ligand is generated at an appropriate level and at a defined time and space during embryonic development. Our major goal is to understand how the proteases required to produce the Toll ligand are organized into a pathway and how they are activated and regulated.
Our specific aims are: 1) To test if the proteases act in a sequential activation cascade and if the activation of any of the proteases is spatially regulated, by biochemica1 analysis of the proteases in vivo; 2) To determine if the proteases are activated on the embryo surface and if activation is spatially restricted, using immunolocalization methods; 3) To decipher the role of the GD protein in the pathway, through a combination of molecular genetic and biochemical experiments; and 4) To identify factors that spatially control the activity of the Snake and Easter proteases, using biochemical and genetic approaches. Recent studies suggest that the ancestral role of the Toll signaling pathway is in host defense, to recognize and signal the presence of microbial pathogens, and is shared by the immune systems of insects and mammals. The signal that acts as the ligand for Toll in mammalian immunity, or how this signal is generated, is not known. Thus, the information obtained from our proposed studies should be of significant relevance for addressing the question of how the Toll signaling pathway is activated during the immune response.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM049370-11
Application #
6636084
Study Section
Cell Development and Function Integrated Review Group (CDF)
Program Officer
Greenberg, Judith H
Project Start
1992-08-01
Project End
2004-12-14
Budget Start
2003-04-01
Budget End
2004-12-14
Support Year
11
Fiscal Year
2003
Total Cost
$326,681
Indirect Cost
Name
Yale University
Department
Anatomy/Cell Biology
Type
Schools of Medicine
DUNS #
043207562
City
New Haven
State
CT
Country
United States
Zip Code
06520
Hyun, Joogyung; Hashimoto, Carl (2011) Physiological effects of manipulating the level of insulin-degrading enzyme in insulin-producing cells of Drosophila. Fly (Austin) 5:53-7
Tang, Huaping (2009) Regulation and function of the melanization reaction in Drosophila. Fly (Austin) 3:105-11
Scherfer, Christoph; Tang, Huaping; Kambris, Zakaria et al. (2008) Drosophila Serpin-28D regulates hemolymph phenoloxidase activity and adult pigmentation. Dev Biol 323:189-96
Tang, Huaping; Kambris, Zakaria; Lemaitre, Bruno et al. (2008) A serpin that regulates immune melanization in the respiratory system of Drosophila. Dev Cell 15:617-26
Jang, In-Hwan; Chosa, Naoyuki; Kim, Sung-Hee et al. (2006) A Spatzle-processing enzyme required for toll signaling activation in Drosophila innate immunity. Dev Cell 10:45-55
Tang, Huaping; Kambris, Zakaria; Lemaitre, Bruno et al. (2006) Two proteases defining a melanization cascade in the immune system of Drosophila. J Biol Chem 281:28097-104
Richer, Martin J; Juliano, Luiz; Hashimoto, Carl et al. (2004) Serpin mechanism of hepatitis C virus nonstructural 3 (NS3) protease inhibition: induced fit as a mechanism for narrow specificity. J Biol Chem 279:10222-7
Richer, Martin J; Keays, Clairessa A; Waterhouse, Jennifer et al. (2004) The Spn4 gene of Drosophila encodes a potent furin-directed secretory pathway serpin. Proc Natl Acad Sci U S A 101:10560-5
Hashimoto, Carl; Kim, Dong Ryoung; Weiss, Linnea A et al. (2003) Spatial regulation of developmental signaling by a serpin. Dev Cell 5:945-50
Turcotte, Cynthia L; Hashimoto, Carl (2002) Evidence for a glycosaminoglycan on the nudel protein important for dorsoventral patterning of the drosophila embryo. Dev Dyn 224:51-7

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