title: Ectopic overexpression of c-mpl by retroviral-mediated gene transfer suppressed megakaryopoiesis but enhanced erythropoiesis in mice.
abstract: In this report, we tested whether ectopic overexpression of a cell surface receptor cDNA could be used to explore the physiological roles of that receptor. We generated c-mpl overexpressing animals by reconstituting mice with retroviral vector-transduced bone marrow (BM) cells. We observed that platelet counts in the c-mpl overexpressing mice failed to recover to normal levels and remained at <200 x 10(6)/mL post-transplantation, while platelet numbers in the control mice returned to > 800 x 10(6)/mL by 4 weeks post-transplantation. However, platelet counts in the c-mpl overexpressing mice could be stimulated to normal levels after administration of rhMGDF. No significant changes in peripheral leukocyte counts were observed, although the number of CFU-E, GM-CFC, and CFC-multi were reduced two- to threefold in the BM of the c-mpl overexpressing mice. In addition, enhanced erythropoiesis was observed in the c-mpl overexpressing mice. The mpl receptors on erythroid cells were functional as demonstrated by tyrosine-phosphorylation of mpl receptor on RBC and by in vitro erythroid colony-formation in response to MGDF stimulation, respectively. These results suggested that ectopically expressed mpl receptors competed for ligand in vivo leading to an insufficient amount of circulating thrombopoietin (Tpo) for the development of megakaryocytic lineage. These results further suggest that, in addition to sequestering circulating Tpo, overexpression of the mpl receptor on erythroid progenitors may directly contribute to enhanced erythropoiesis in vivo. Our studies demonstrate that ectopic overexpression of a receptor by retroviral-mediated gene transfer provides an approach to explore the biological roles of novel receptors.
authors.affiliation: Department of Pathology, Amgen Inc., Thousand Oaks, CA 91320-1789, USA. XYan@amgen.com
authors.forename: X Q
authors.initials: XQ
authors.lastname: Yan
authors.forename: D L
authors.initials: DL
authors.lastname: Lacey
authors.forename: C
authors.initials: C
authors.lastname: Saris
authors.forename: S
authors.initials: S
authors.lastname: Mu
authors.forename: D
authors.initials: D
authors.lastname: Hill
authors.forename: R G
authors.initials: RG
authors.lastname: Hawley
authors.forename: F A
authors.initials: FA
authors.lastname: Fletcher
mesh_terms: D000818:Animals; D003114:Colony-Forming Units Assay; D018076:DNA, Complementary / Q000235:genetics; D004920:Erythropoiesis; D005822:Genetic Vectors / Q000235:genetics; D006410:Hematopoiesis; D016298:Hematopoietic Cell Growth Factors / Q000494:pharmacology; D018380:Hematopoietic Stem Cell Transplantation; D006412:Hematopoietic Stem Cells / Q000821:virology; D007958:Leukocyte Count; D008533:Megakaryocytes / Q000166:cytology*; D051379:Mice; D008810:Mice, Inbred C57BL; D008811:Mice, Inbred DBA; D009363:Neoplasm Proteins; D011518:Proto-Oncogene Proteins / Q000096:biosynthesis* / Q000235:genetics; D011533:Proviruses / Q000235:genetics; D018121:Receptors, Cytokine; D053628:Receptors, Thrombopoietin; D011993:Recombinant Fusion Proteins / Q000494:pharmacology / Q000502:physiology; D011994:Recombinant Proteins / Q000494:pharmacology; D012190:Retroviridae / Q000235:genetics; D013926:Thrombopoietin / Q000494:pharmacology / Q000502:physiology; D014162:Transfection
chemical_list: D018076:DNA, Complementary; D016298:Hematopoietic Cell Growth Factors; D009363:Neoplasm Proteins; D011518:Proto-Oncogene Proteins; D018121:Receptors, Cytokine; D053628:Receptors, Thrombopoietin; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; C082218:MPL protein, human; D013926:Thrombopoietin