Interleukin-7 (IL-7) is an essential pleiotrophic cytokine regulating numerous immunological events. Normal IL-7 signaling responses trigger the development, proliferation, and homeostasis of T and B cells as well as remodeling of the extracellular matrix. IL-7 activates its signaling cascade by interacting with its own cell surface receptor, IL-7R alpha, and the common gamma chain receptor through a cytokine-induced receptor heterodimerization mechanism. Under- and over-stimulation of the IL-7 pathway has been implicated in the pathogenesis of a form of severe combined immunodeficiency, autoimmune conditions, coronary artery disease, and several cancers. Structural, energetic, and cell biological studies of IL-7-induced receptor heterodimerization require a careful analysis of two distinct events: the association of IL-7 to IL-7R alpha and of IL-7:IL-7R alpha complex to the common gamma chain receptor. This research proposal will focus on the initiation step, the binding interaction of IL-7 with its alpha receptor. Results show that glycosylation of the IL- 7R alpha confers optimal binding affinity to IL-7. The importance of glycosylation of the IL-7R alpha appears to be a novel binding recognition mechanism for the common gamma chain receptor family. The thermodynamic and kinetic properties of the wild-type IL-7:IL-7R alpha interaction will be ascertained using a variety of biophysical methods including isothermal titration calorimetry and surface plasmon resonance. The functional epitopes on both IL-7 and IL-7R alpha will be determined using site-directed scanning mutagenesis, biophysical methods, and cell proliferation assays. Three-dimensional structures of free IL-7, free unglycosylated and glycosylated IL-7R alpha, and complexes of IL-7 bound to unglycosylated and glycosylated IL-7R alpha will be determined using x-ray crystallography or NMR spectroscopy. Our understanding of the binding determinants for the IL-7-receptor interaction will be used to engineer novel peptide and protein antagonists using rational protein design and phage display techniques. The long-term goal of this research involves development of a structural energetic model to explain the normal and aberrant IL-7 signaling mechanisms and the generation of new therapeutic peptides/proteins to treat the many deficiencies and diseases associated with impaired IL-7 signaling. Public Health Relevance Statement: This research proposal will focus on the structural, energetic, and cell biological studies of the interaction between human interleukin-7 and its receptor, interleukin-7 receptor alpha. Our understanding of the binding determinants for the IL-7-receptor interaction will be used to engineer novel peptide and protein antagonists using rational protein design and phage display techniques. The long-term goal of this research involves development of a structural energetic model to explain the normal and aberrant IL-7 signaling mechanisms and the generation of new therapeutic peptides/proteins to treat the many deficiencies and diseases associated with impaired human IL-7 signaling.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI072142-07
Application #
8206719
Study Section
Cellular and Molecular Immunology - A Study Section (CMIA)
Program Officer
Leitner, Wolfgang W
Project Start
2008-01-15
Project End
2013-12-31
Budget Start
2012-01-01
Budget End
2013-12-31
Support Year
7
Fiscal Year
2012
Total Cost
$294,030
Indirect Cost
$98,010
Name
University of Maryland College Park
Department
Miscellaneous
Type
Other Domestic Higher Education
DUNS #
790934285
City
College Park
State
MD
Country
United States
Zip Code
20742
Lundström, Wangko; Highfill, Steven; Walsh, Scott T R et al. (2013) Soluble IL7R? potentiates IL-7 bioactivity and promotes autoimmunity. Proc Natl Acad Sci U S A 110:E1761-70
Zhang, Fuming; Liang, Xinle; Pu, Dennis et al. (2012) Biophysical characterization of glycosaminoglycan-IL-7 interactions using SPR. Biochimie 94:242-9
McElroy, Craig A; Holland, Paul J; Zhao, Peng et al. (2012) Structural reorganization of the interleukin-7 signaling complex. Proc Natl Acad Sci U S A 109:2503-8
Singh, Arpita; Del Poeta, Maurizio (2011) Lipid signalling in pathogenic fungi. Cell Microbiol 13:177-85
Williams, Virginia; Del Poeta, Maurizio (2011) Role of glucose in the expression of Cryptococcus neoformans antiphagocytic protein 1, App1. Eukaryot Cell 10:293-301
Garcia, Jacqueline; Sims, Kellie J; Schwacke, John H et al. (2011) Biochemical systems analysis of signaling pathways to understand fungal pathogenicity. Methods Mol Biol 734:173-200
Rhome, Ryan; Singh, Arpita; Kechichian, Talar et al. (2011) Surface localization of glucosylceramide during Cryptococcus neoformans infection allows targeting as a potential antifungal. PLoS One 6:e15572
Singh, Arpita; Qureshi, Asfia; Del Poeta, Maurizio (2011) Quantitation of cellular components in Cryptococcus neoformans for system biology analysis. Methods Mol Biol 734:317-33
Walsh, Scott T R (2010) A biosensor study indicating that entropy, electrostatics, and receptor glycosylation drive the binding interaction between interleukin-7 and its receptor. Biochemistry 49:8766-78
McQuiston, Travis; Luberto, Chiara; Del Poeta, Maurizio (2010) Role of host sphingosine kinase 1 in the lung response against Cryptococcosis. Infect Immun 78:2342-52

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