title: The erythropoietin receptor transmembrane domain mediates complex formation with viral anemic and polycythemic gp55 proteins.
abstract: Erythropoietin receptor (EpoR) activation is crucial for mature red blood cell production. The murine EpoR can also be activated by the envelope protein of the polycythemic (P) spleen focus forming virus (SFFV), gp55-P. Due to differences in the TM sequence, gp55 of the anemic (A) strain SFFV, gp55-A, cannot efficiently activate the EpoR. Using antibody-mediated immunofluorescence co-patching, we show that the majority of EpoR forms hetero-oligomers at the cell surface with gp55-P and, surprisingly, with gp55-A. The EpoR TM domain is targeted by gp55-P and -A, as only chimeric receptors containing EpoR TM sequences oligomerized with gp55 proteins. Both gp55-P and gp55-A are homodimers on the cell surface, as shown by co-patching. However, when the homomeric interactions of the isolated TM domains were assayed by TOXCAT bacterial reporter system, only the TM sequence of gp55-P was dimerized. Thus, homo-oligomerization of gp55 proteins is insufficient for full EpoR activation, and a correct conformation of the dimer in the TM region is required. This is supported by the failure of gp55-A-->P, a mutant protein whose TM domain can homo-oligomerize, to fully activate EpoR. As unliganded EpoR forms TM-dependent but inactive homodimers, we propose that the EpoR can be activated to different extents by homodimeric gp55 proteins, depending on the conformation of the gp55 protein dimer in the TM region.
authors.affiliation: Ludwig Institute for Cancer Research and University of Louvain, Brussels B-1200, Belgium. stefan.constantinescu@bru.licr.org
authors.forename: Stefan N
authors.initials: SN
authors.lastname: Constantinescu
authors.forename: Tzvia
authors.initials: T
authors.lastname: Keren
authors.forename: William P
authors.initials: WP
authors.lastname: Russ
authors.forename: Iban
authors.initials: I
authors.lastname: Ubarretxena-Belandia
authors.forename: Yaniv
authors.initials: Y
authors.lastname: Malka
authors.forename: Katharina F
authors.initials: KF
authors.lastname: Kubatzky
authors.forename: Donald M
authors.initials: DM
authors.lastname: Engelman
authors.forename: Harvey F
authors.initials: HF
authors.lastname: Lodish
authors.forename: Yoav I
authors.initials: YI
authors.lastname: Henis
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies / Q000737:chemistry; D015153:Blotting, Western; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism*; D019281:Dimerization; D000939:Epitopes; D017930:Genes, Reporter; D051379:Mice; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D009154:Mutation; D010957:Plasmids / Q000378:metabolism; D011233:Precipitin Tests; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D051381:Rats; D017467:Receptors, Erythropoietin / Q000737:chemistry*; D017386:Sequence Homology, Amino Acid; D014162:Transfection; D014759:Viral Envelope Proteins / Q000737:chemistry* / Q000378:metabolism
chemical_list: D000906:Antibodies; D000939:Epitopes; D017467:Receptors, Erythropoietin; D014759:Viral Envelope Proteins; C078780:glycoprotein gp55, Friend spleen focus-forming virus