Megakaryocytopoietic activity of a truncated variant of mouse thrombopoietin.
Thrombopoietin (TPO) is a hemopoietic cytokine that specifically stimulates the growth and development of megakaryocytes. In addition to full-length TPO (corresponding to TPO-1), cDNAs of three truncated variants (TPO-2 to 4) have been isolated. However, whether or not these variants have biological activities has not been clarified presumably because of poor secretion into extracellular milieu. To examine the biological significance of the shortest variants, TPO-4, we prepared two fusion cDNAs of TPO-1 and TPO-4, each of which was combined with the finger-growth factor-kringle 1 (FGK1) domains of tissue plasminogen activator (tPA) as a tag and transfected them into baby hamster kidney (BHK) cells. An immunodetection assay for FGK1 antigen revealed that the amount of secreted TPO-4-FGK1 fusion protein was about 2% of that of TPO-1-FGK1. These fusion proteins after affinity purification had comparable activities on a molar basis in both megakaryocyte colony stimulating activity towards mouse bone marrow cells and growth promotion activity towards the TPO-responsive cell line UT-7/GM. These results indicate that the TPO-4 is biologically active in vitro, although its primary structure of the C-terminal half after the 111th amino acid residues is completely different from authentic TPO-1 due to frame shift.
Hoshi
S
S
Tsukuba Research Laboratories, Eisai Co., Ltd., Ibaraki, Japan.
Yoshitomi
H
H
Komatsu
N
N
Yoshitake
S
S
Okada
M
M
D000818:Animals; D001854:Bone Marrow Cells; D002460:Cell Line; D006224:Cricetinae; D006410:Hematopoiesis / Q000187:drug effects*; D018082:Kringles; D008533:Megakaryocytes / Q000166:cytology*; D051379:Mice; D011993:Recombinant Fusion Proteins; D013329:Structure-Activity Relationship; D013926:Thrombopoietin / Q000737:chemistry* / Q000494:pharmacology; D010959:Tissue Plasminogen Activator / Q000737:chemistry
D011993:Recombinant Fusion Proteins; D013926:Thrombopoietin; D010959:Tissue Plasminogen Activator