Pasteurella multocida is a pathogenic bacterium associated with the agricultural diseases pasteurellosis, hemorrhagic septicemia, dermonecrosis, and progressive atrophic rhinitis. P. multocida can cause severe complications in human infections from animal bites or scratches, respiratory infections, and exposure to animals during pregnancy. P. multocida toxin is an important virulence factor of P. multocida, and purified PMT alone is sufficient to experimentally induce progressive atrophic rhinitis. PMT appears to enter cells via receptor-mediated endocytosis and causes activation of signal transduction events and DNA synthesis. Recent studies from our laboratory have identified G alpha protein as the primary target of PMT action that activates the phosphatidylinositol-specific phospholipase C-beta 1 and the inositol triphosphate pathway in Xenopus oocytes. Studies from our laboratory have also shown that the N-terminus of PMT is important for this activity, and we have proposed a model for PMT's intracellular action. We have cloned the entire toxA gene (1285 residues) from P. multocida and have generated a number of deletion mutants, encoding residues 1-73, 1-293, 1-506, 506- 1285, and 1059-1285. Our long range goals are to use these recombinant proteins to understand the structure and mechanism of action of PMT at the molecular and biochemical level, both to facilitate future therapeutic intervention in the bacterial pathogenesis of P. multocida , as well as to provide insight into the molecular signalling events involved in the control of cell growth and differentiation. In particular, we hope to demonstrate the utility of PMT as a new biochemical tool for studying intracellular signalling pathways involving the Gq family of regulatory proteins. To achieve our goals, we propose the following: (1) To define the functional domains of the protein, so as to determine which of the toxin's domains are responsible for (1) binding to the eukaryotic cell receptors and (2) stimulating the intracellular signal transduction pathways. (2) To elucidate the molecular mechanism by which PMT activates Gq- protein, by determining whether PMT's activation of Gq-protein is caused by a covalent modification or by noncovalent interaction. (3) To test the hypothesis that PMT uncouples the ligand-regulated interaction between receptor and Gq-protein, using the Xenopus oocyte system overexpressing exogenous 5-HT2 receptor and Gqalpha-protein.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
First Independent Research Support & Transition (FIRST) Awards (R29)
Project #
7R29AI038396-05
Application #
6263280
Study Section
Bacteriology and Mycology Subcommittee 2 (BM)
Program Officer
Baker, Phillip J
Project Start
1996-08-01
Project End
2001-07-31
Budget Start
1999-09-01
Budget End
2000-07-31
Support Year
5
Fiscal Year
1999
Total Cost
Indirect Cost
Name
University of Illinois Urbana-Champaign
Department
Microbiology/Immun/Virology
Type
Schools of Arts and Sciences
DUNS #
041544081
City
Champaign
State
IL
Country
United States
Zip Code
61820
Brothers, Michael C; Geissler, Brett; Hisao, Grant S et al. (2014) Backbone and side-chain resonance assignments of the membrane localization domain from Pasteurella multocida toxin. Biomol NMR Assign 8:221-4
Brothers, Michael C; Geissler, Brett; Hisao, Grant S et al. (2014) Backbone and side-chain assignments of an effector membrane localization domain from Vibrio vulnificus MARTX toxin. Biomol NMR Assign 8:225-8
Wilson, Brenda A; Ho, Mengfei (2013) Pasteurella multocida: from zoonosis to cellular microbiology. Clin Microbiol Rev 26:631-55
Oubrahim, Hammou; Wong, Allison; Wilson, Brenda A et al. (2013) Mammalian target of rapamycin complex 1 (mTORC1) plays a role in Pasteurella multocida toxin (PMT)-induced protein synthesis and proliferation in Swiss 3T3 cells. J Biol Chem 288:2805-15
Gargi, Amandeep; Tamilselvam, Batcha; Powers, Brendan et al. (2013) Cellular interactions of the cytolethal distending toxins from Escherichia coli and Haemophilus ducreyi. J Biol Chem 288:7492-505
Bannai, Yuka; Aminova, Leila R; Faulkner, Melinda J et al. (2012) Rho/ROCK-dependent inhibition of 3T3-L1 adipogenesis by G-protein-deamidating dermonecrotic toxins: differential regulation of Notch1, Pref1/Dlk1, and ?-catenin signaling. Front Cell Infect Microbiol 2:80
Wilson, Brenda A; Ho, Mengfei (2012) Pasteurella multocida toxin interaction with host cells: entry and cellular effects. Curr Top Microbiol Immunol 361:93-111
Brothers, Michael C; Ho, Mengfei; Maharjan, Ram et al. (2011) Membrane interaction of Pasteurella multocida toxin involves sphingomyelin. FEBS J 278:4633-48
Wilson, Brenda A; Ho, Mengfei (2011) Cellular and molecular action of the mitogenic protein-deamidating toxin from Pasteurella multocida. FEBS J 278:4616-32
Wilson, Brenda A; Ho, Mengfei (2010) Recent insights into Pasteurella multocida toxin and other G-protein-modulating bacterial toxins. Future Microbiol 5:1185-201

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