Fusion of the erythropoietin receptor and the Friend spleen focus-forming virus gp55 glycoprotein transforms a factor-dependent hematopoietic cell line.
The Friend spleen focus-forming virus (SFFV) gp55 glycoprotein binds to the erythropoietin receptor (EPO-R), causing constitutive receptor signaling and the first stage of Friend erythroleukemia. We have used three independent strategies to further define this transforming molecular interaction. First, using a retroviral selection strategy, we have isolated the cDNAs encoding three fusion polypeptides containing regions of both EPO-R and gp55. These fusion proteins, like full-length gp55, transformed the Ba/F3 factor-dependent hematopoietic cell line and localized the transforming activity of gp55 to its transmembrane domain. Second, we have isolated a mutant of gp55 (F-gp55-M1) which binds, but fails to activate, EPO-R. We have compared the transforming activity of this gp55 mutant with the EPO-R-gp55 fusion proteins and with other variants of gp55, including wild-type polycythemia Friend gp55 and Rauscher gp55. All of the fusion polypeptides and mutant gp55 polypeptides were expressed at comparable levels, and all coimmunoprecipitated with wild-type EPO-R, but only the Friend gp55 and the EPO-R-gp55 fusion proteins constitutively activated wild-type EPO-R. Third, we have examined the specificity of the EPO-R-gp55 interaction by comparing the differential activation of murine and human EPO-R by gp55. Wild-type gp55 had a highly specific interaction with murine EPO-R; gp55 bound, but did not activate, human EPO-R.
Showers
M O
MO
Division of Pediatric Oncology, Dana-Farber Cancer Institute, Harvard Medical School, Boston, Massachusetts 02115.
DeMartino
J C
JC
Saito
Y
Y
D'Andrea
A D
AD
D000818:Animals; D002460:Cell Line; D002472:Cell Transformation, Viral; D003001:Cloning, Molecular; D005091:Exons; D005622:Friend murine leukemia virus / Q000378:metabolism*; D006412:Hematopoietic Stem Cells; D006801:Humans; D051379:Mice; D009154:Mutation; D010957:Plasmids; D011485:Protein Binding; D017467:Receptors, Erythropoietin / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D013045:Species Specificity; D014759:Viral Envelope Proteins / Q000235:genetics / Q000378:metabolism*
D017467:Receptors, Erythropoietin; D011993:Recombinant Fusion Proteins; D014759:Viral Envelope Proteins; C078780:glycoprotein gp55, Friend spleen focus-forming virus