Entamoeba histolytica is a human cytolytic parasite with worldwide distribution that infects up to 50 million people and results in an estimated 70,000 deaths annually. The E. histolytica Ga1NAc lectin mediates trophozoite adherence and plays a role in contact-dependent killing of human immune effector and epithelial cells. The cytolytic capacity of E. histolytica is central to pathogenesis yet little is known about how cytolysis is initiated after cell contact via the Ga1NAc lectin. The hypothesis of this proposal is that protein tyrosine kinases and/or their substrates are intimately involved in the signal transduction by the Ga1NAc lectin. Protein tyrosine kinases (PTK) and/or their substrates have been demonstrated by co-immunoprecipitation to associate with the Ga1NAc lectin. We propose to test the role of these PTK/substrates in lectin- mediated signal transduction. Ga1NAc lectin-associated PTK/substrates will be identified from lectin co-immunoprecipitations or by the yeast two hybrid system (using the lectin cytoplasmic domain as bait). Lectin-associated PTK/substrates will be cloned and sequenced, the primary amino acid sequence characterized, and mAb produced against them. These reagents will be used to assess changes in the PTK/substrate activity, phosphorylation state and intracellular location during signaling. Finally genetic characterization will be attempted using reverse genetic approaches. Successful completion of these experiments will provide insight into the role and identity of PTK/substrates in lectin-mediated killing and may establish the basis for novel therapeutic or prophylactic measures against amebiasis.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
2R01AI032615-05
Application #
2468580
Study Section
Tropical Medicine and Parasitology Study Section (TMP)
Project Start
1993-12-01
Project End
2002-11-30
Budget Start
1997-12-01
Budget End
1998-11-30
Support Year
5
Fiscal Year
1998
Total Cost
Indirect Cost
Name
University of Virginia
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
001910777
City
Charlottesville
State
VA
Country
United States
Zip Code
22904
Okada, Mami; Huston, Christopher D; Oue, Miho et al. (2006) Kinetics and strain variation of phagosome proteins of Entamoeba histolytica by proteomic analysis. Mol Biochem Parasitol 145:171-83
McCoy, James J; Mann, Barbara J (2005) Proteomic analysis of Gal/GalNAc lectin-associated proteins in Entamoeba histolytica. Exp Parasitol 110:220-5
Okada, Mami; Huston, Christopher D; Mann, Barbara J et al. (2005) Proteomic analysis of phagocytosis in the enteric protozoan parasite Entamoeba histolytica. Eukaryot Cell 4:827-31
Hughes, Molly A; Lee, Constance W; Holm, Christopher F et al. (2003) Identification of Entamoeba histolytica thiol-specific antioxidant as a GalNAc lectin-associated protein. Mol Biochem Parasitol 127:113-20
Teixeira, Jose E; Mann, Barbara J (2002) Entamoeba histolytica-induced dephosphorylation in host cells. Infect Immun 70:1816-23
Petri Jr, William A; Haque, Rashidul; Mann, Barbara J (2002) The bittersweet interface of parasite and host: lectin-carbohydrate interactions during human invasion by the parasite Entamoeba histolytica. Annu Rev Microbiol 56:39-64
Godbold, Gene D; Corbett, Kevin D; Mann, Barbara J (2002) A Rho-like small GTPase of Entamoeba histolytica contains an unusual amino acid residue in a conserved GDP-stabilization region and is not a substrate for C3 exoenzyme. Exp Parasitol 101:107-10
Cheng, X J; Hughes, M A; Huston, C D et al. (2001) Intermediate subunit of the Gal/GalNAc lectin of Entamoeba histolytica is a member of a gene family containing multiple CXXC sequence motifs. Infect Immun 69:5892-8
Godbold, G D; Mann, B J (2000) Cell killing by the human parasite Entamoeba histolytica is inhibited by the rho-inactivating C3 exoenzyme. Mol Biochem Parasitol 108:147-51
Hughes, M A; Reed, S L; Mann, B J (2000) Role of the Entamoeba histolytica Gal/GalNAc lectin in recruiting proteins to the host:parasite interface. Arch Med Res 31:S229-30

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