title: Association of nucleophosmin negatively regulates CXCR4-mediated G protein activation and chemotaxis.
abstract: CXCR4, the primary receptor for CXCL12, plays a critical role in the development of hematopoietic, vascular, central nervous, and immune systems by mediating directional migration of precursor cells. This mechanism promotes homing of tumor cells to metastatic sites that secrete CXCL12, and CXCR4 expression is a negative prognostic factor in acute myelogenous leukemia (AML). To elucidate mechanisms that regulate CXCR4 signaling, we used a proteomic approach to identify proteins physically associated with CXCR4. Analysis of CXCR4 immune complexes identified nucleophosmin (NPM), which was confirmed by reciprocal coimmunoprecipitation for NPM. Constitutively active CXCR4 variants bound higher levels of NPM than the wild-type receptor, which was reversed by T140, an inverse agonist. NPM binding to CXCR4 localized interactions to the C terminus and cytoplasmic loop (CL)-3, but not CL-1 or CL-2. Alanine scanning mutagenesis demonstrated that positively charged amino acids in CL-3 were critical for NPM binding. Recombinant NPM decreased GTP binding in membrane fractions after activation of CXCR4 by CXCL12. Suppression of NPM expression enhanced chemotactic responses to CXCL12, and, conversely, overexpression of a cytosolic NPM mutant reduced chemotaxis induced by CXCL12. This study provides evidence for a novel role for NPM as a negative regulator of CXCR4 signaling induced by CXCL12 that may be relevant to the biology of AML.
authors: Department of Pathology, Medical College of Georgia, Augusta, GA 30912, USA.
authors: Wenbo
authors: W
authors: Zhang
authors: Jean-Marc
authors: JM
authors: Navenot
authors: Nicole M
authors: NM
authors: Frilot
authors: Nobutaka
authors: N
authors: Fujii
authors: Stephen C
authors: SC
authors: Peiper
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D002633:Chemotaxis; D003600:Cytosol / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000378:metabolism*; D006801:Humans; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000737:chemistry / Q000378:metabolism*; D000090243:Nucleophosmin; D011487:Protein Conformation; D054730:Protein Interaction Domains and Motifs; D025941:Protein Interaction Mapping; D019718:Receptors, CXCR4 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: C000716967:NPM1 protein, human; D009687:Nuclear Proteins; D019718:Receptors, CXCR4; D000090243:Nucleophosmin; D019204:GTP-Binding Proteins