The goal of this project is to understand the structural and functional requirements for chemokine signaling in vivo. Chemokines are small secreted proteins that mediate inflammation, stem cell homing and early embryonic development by providing directional cues to migrating cells. The 50 known chemokines induce chemotaxis by specifically activating members of a group of 20 G protein-coupled receptors, which are integral membrane proteins of the seven transmembrane type. A chemokine concentration gradient maintained by interactions with extracellular matrix glycosaminoglycans (GAG) is also needed to induce chemotactic responses in vivo, and, for some chemokines, formation of homodimeric structures is also a functional requirement. Stromal cell-derived factor-1 (SDF1) and its cognate receptor CXCR4 comprise a chemokine signaling system that is exploited by metastatic cancers and HIV/AIDS. HIV-1 gains entry to T cells through specific binding to CXCR4, a process that is inhibited by the chemokine SDF1. Cancer cells that express CXCR4 and escape the primary tumor environment travel the circulatory and lymphatic systems homing in on a select group of tissues that constitutively produce high levels of SDF1, including bone marrow, lung and lymph nodes. A complete model for chemokine function at a molecular level will include GPCR activation, GAG binding and dimer formation. We intend to combine NMR spectroscopy with mutagenesis studies and functional assays to characterize each of the specific binding interactions required for SDF1-CXCR4 signaling.
In Aim 1, we will determine whether SDF1 functions as a monomer or dimer by designing mutants that form only one or the other species and testing their activities in vivo. Experiments in Aim 2 will determine the structure of an SDF1-GAG complex, and explore the relationship between GAG binding and dimerization.
In Aim 3 we will determine the structure of a complex between SDF1 and a soluble fragment of CXCR4, and pursue NMR studies of the full-length CXCR4 protein solubilized in detergent micelles. These studies will enable the construction of a unified model for SDF1 activity in vivo that accounts for all its known functional interactions.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI058072-03
Application #
7012754
Study Section
Physical Biochemistry Study Section (PB)
Program Officer
Nasseri, M Faraz
Project Start
2004-08-01
Project End
2009-01-31
Budget Start
2006-02-01
Budget End
2007-01-31
Support Year
3
Fiscal Year
2006
Total Cost
$258,895
Indirect Cost
Name
Medical College of Wisconsin
Department
Biochemistry
Type
Schools of Medicine
DUNS #
937639060
City
Milwaukee
State
WI
Country
United States
Zip Code
53226
Gao, Xianlong; Albee, Lauren J; Volkman, Brian F et al. (2018) Asymmetrical ligand-induced cross-regulation of chemokine (C-X-C motif) receptor 4 by ?1-adrenergic receptors at the heteromeric receptor complex. Sci Rep 8:2730
Szpakowska, Martyna; Nevins, Amanda M; Meyrath, Max et al. (2018) Different contributions of chemokine N-terminal features attest to a different ligand binding mode and a bias towards activation of ACKR3/CXCR7 compared with CXCR4 and CXCR3. Br J Pharmacol 175:1419-1438
Albee, Lauren J; LaPorte, Heather M; Gao, Xianlong et al. (2018) Identification and functional characterization of arginine vasopressin receptor 1A : atypical chemokine receptor 3 heteromers in vascular smooth muscle. Open Biol 8:
Egner, John M; Jensen, Davin R; Olp, Michael D et al. (2018) Development and Validation of 2D Difference Intensity Analysis for Chemical Library Screening by Protein-Detected NMR Spectroscopy. Chembiochem 19:448-458
Dishman, Acacia F; Volkman, Brian F (2018) Unfolding the Mysteries of Protein Metamorphosis. ACS Chem Biol 13:1438-1446
Thomas, Monica A; Kleist, Andrew B; Volkman, Brian F (2018) Decoding the chemotactic signal. J Leukoc Biol 104:359-374
Hall, Cherisse L; Lytle, Betsy L; Jensen, Davin et al. (2017) Structure and Dimerization of IreB, a Negative Regulator of Cephalosporin Resistance in Enterococcus faecalis. J Mol Biol 429:2324-2336
Albee, Lauren J; Eby, Jonathan M; Tripathi, Abhishek et al. (2017) ?1-Adrenergic Receptors Function Within Hetero-Oligomeric Complexes With Atypical Chemokine Receptor 3 and Chemokine (C-X-C motif) Receptor 4 in Vascular Smooth Muscle Cells. J Am Heart Assoc 6:
Getschman, A E; Imai, Y; Larsen, O et al. (2017) Protein engineering of the chemokine CCL20 prevents psoriasiform dermatitis in an IL-23-dependent murine model. Proc Natl Acad Sci U S A 114:12460-12465
Cheng, You-Hong; Eby, Jonathan M; LaPorte, Heather M et al. (2017) Effects of cognate, non-cognate and synthetic CXCR4 and ACKR3 ligands on human lung endothelial cell barrier function. PLoS One 12:e0187949

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