Stimulating cell proliferation through the pharmacologic activation of c-kit.
Previous studies have shown that expression of a membrane targeted chimeric protein containing the erythropoietin receptor (EpoR) cytoplasmic domain fused to the FK506-binding peptide FKBP12 allowed Ba/F3 cells to be rescued from interleukin-3 (IL-3) deprivation using a dimeric form of FK506, called FK1012. In this report, a similar approach is applied to the c-kit receptor. Expression of a membrane targeted fusion protein containing the c-kit receptor linked to one or more copies of FKBP12 allowed Ba/F3 cells to be switched from IL-3 dependence to FK1012-dependence. Similar results were obtained using an alternative dimerizer of FKBP12 domains called AP1510. Pharmacologic dimerization of chimeric proteins containing only a single FKBP12 domain confirmed that receptor dimerization is sufficient for proliferative signaling. Interestingly, while the proliferative effects of both FK1012 and AP1510 were reversible, FK1012-driven proliferation persisted for several days after drug withdrawal. Furthermore, much higher concentrations of FK506 were required to inhibit FK1012-mediated proliferation than were required to inhibit AP1510-mediated proliferation. The persistence of FK1012's effect appeared to be specific to clones expressing c-kit-containing fusion proteins. These results suggest that pharmacologically-responsive fusion proteins containing c-kit may be useful for specifically and reversibly expanding genetically modified hematopoietic cell populations.
Jin
L
L
Department of Medicine, University of Washington, Seattle 98195, USA.
Asano
H
H
Blau
C A
CA
D000818:Animals; D002352:Carrier Proteins / Q000737:chemistry / Q000235:genetics / Q000502:physiology; D002455:Cell Division / Q000187:drug effects; D002460:Cell Line; D002462:Cell Membrane / Q000737:chemistry; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000502:physiology; D019281:Dimerization; D015870:Gene Expression; D006360:Heat-Shock Proteins / Q000737:chemistry / Q000235:genetics / Q000502:physiology; D006412:Hematopoietic Stem Cells / Q000166:cytology / Q000378:metabolism; D007377:Interleukin-3 / Q000494:pharmacology; D051379:Mice; D019009:Proto-Oncogene Proteins c-kit / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D011993:Recombinant Fusion Proteins; D015398:Signal Transduction; D016559:Tacrolimus / Q000031:analogs & derivatives / Q000494:pharmacology; D022021:Tacrolimus Binding Proteins; D014162:Transfection
D002352:Carrier Proteins; D004268:DNA-Binding Proteins; C084260:FK 1012; D006360:Heat-Shock Proteins; D007377:Interleukin-3; D011993:Recombinant Fusion Proteins; D019009:Proto-Oncogene Proteins c-kit; D022021:Tacrolimus Binding Proteins; D016559:Tacrolimus