title: Modulation of the CXC chemokine receptor 4 agonist activity of ubiquitin through C-terminal protein modification.
abstract: Extracellular ubiquitin has recently been described as a CXC chemokine receptor (CXCR) 4 agonist. Studies on the structure-function relationship suggested that the C-terminus of ubiquitin facilitates CXCR4 activation. It remains unknown, however, whether C-terminal processing of ubiquitin could be biologically relevant and whether modifications of the ubiquitin C-terminus can modulate CXCR4 activation. We show that C-terminal truncated ubiquitin antagonizes ubiquitin and stromal cell-derived factor (SDF)-1α induced effects on cell signaling and function. Reduction of cell surface expression of insulin degrading enzyme (IDE), which cleaves the C-terminal di-Gly of ubiquitin, enhances ubiquitin induced reduction of cAMP levels in BV2 and THP-1 cells, but does not influence changes in cAMP levels in response to SDF-1α. Reduction of cell surface IDE expression in THP-1 cells also increases the chemotactic activity of ubiquitin. As compared with native ubiquitin, C-terminal Tyr extension of ubiquitin results in reduced CXCR4 mediated effects on cellular cAMP levels and abolishes chemotactic activity. Replacement of C-terminal di-Gly of ubiquitin with di-Val or di-Arg enhances CXCR4 mediated effects on cAMP levels and the di-Arg substitution exerts increased chemotactic activity, when compared with wild type ubiquitin. The chemotactic activities of the di-Val and di-Arg mutants and their effects on cAMP levels can be antagonized with C-terminal truncated ubiquitin. These data suggest that the development of CXCR4 ligands with enhanced agonist activities is possible and that C-terminal processing of ubiquitin could constitute a biological mechanism, which regulates termination of receptor signaling.
authors.affiliation: Department of Surgery, Loyola University Chicago, Stritch School of Medicine, Maywood, Illinois 60153, USA.
authors.forename: Abhishek
authors.initials: A
authors.lastname: Tripathi
authors.forename: Vikas
authors.initials: V
authors.lastname: Saini
authors.forename: Adriano
authors.initials: A
authors.lastname: Marchese
authors.forename: Brian F
authors.initials: BF
authors.lastname: Volkman
authors.forename: Wei-Jen
authors.initials: WJ
authors.lastname: Tang
authors.forename: Matthias
authors.initials: M
authors.lastname: Majetschak
mesh_terms: D000818:Animals; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D002469:Cell Separation; D054377:Chemokine CXCL12 / Q000378:metabolism; D000242:Cyclic AMP / Q000378:metabolism; D005434:Flow Cytometry; D020868:Gene Silencing; D006801:Humans; D007328:Insulin / Q000737:chemistry; D051379:Mice; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D034741:RNA, Small Interfering / Q000378:metabolism; D019718:Receptors, CXCR4 / Q000737:chemistry* / Q000378:metabolism; D015398:Signal Transduction; D013329:Structure-Activity Relationship; D014443:Tyrosine / Q000737:chemistry; D025801:Ubiquitin / Q000737:chemistry*
chemical_list: C527276:CXCR4 protein, human; C514936:CXCR4 protein, mouse; D054377:Chemokine CXCL12; D007328:Insulin; D034741:RNA, Small Interfering; D019718:Receptors, CXCR4; D025801:Ubiquitin; D014443:Tyrosine; D000242:Cyclic AMP