Biological activity of human granulocyte colony stimulating factor with a modified C-terminus.
Granulocyte colony-stimulating factor (G-CSF) undergoes receptor-mediated internalization into target cells which are normally restricted to neutrophilic granulocytes and their committed progenitor cells, suggesting that it may be applicable as a myeloid cell-targeting vehicle. To test this notion, we constructed a cDNA encoding a human G-CSF/murine stem cell factor (mSCF) chimeric molecule in a mammalian expression vector and transfected NIH3T3 cells with this plasmid. The resulting chimeric cytokine consisted of the entire G-CSF sequences fused to Lys148 of mSCF. It can be released from the surface membrane of NIH3T3 transformants through proteolytic cleavage at Ala164 of mSCF. The culture media conditioned by a number of stable transformants, which were confirmed by an enzyme-linked immunosorbent assay (ELISA) to secrete an hG-CSF derivative, were examined for their ability to stimulate CFU-G-derived colony formation as well as the proliferation of G-CSF-dependent NFS-60 cells. The results indicated that this C-terminus modified version of hG-CSF is as potent as recombinant hG-CSF in both assays.
Oshima
Y
Y
Department of Hematology, University of Tokyo, Tokyo, 108-8639, Japan.
Tojo
A
A
Niho
Y
Y
Asano
S
S
D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001854:Bone Marrow Cells / Q000166:cytology* / Q000187:drug effects; D002455:Cell Division / Q000187:drug effects; D002460:Cell Line; D002478:Cells, Cultured; D003114:Colony-Forming Units Assay; D017077:Culture Media, Conditioned; D018076:DNA, Complementary; D016179:Granulocyte Colony-Stimulating Factor / Q000096:biosynthesis / Q000737:chemistry* / Q000494:pharmacology*; D006801:Humans; D051379:Mice; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000494:pharmacology; D019089:Stem Cell Factor / Q000096:biosynthesis / Q000737:chemistry / Q000494:pharmacology; D014162:Transfection
D017077:Culture Media, Conditioned; D018076:DNA, Complementary; D011993:Recombinant Fusion Proteins; D019089:Stem Cell Factor; D016179:Granulocyte Colony-Stimulating Factor