Selection of an HEL-derived cell line expressing high levels of platelet factor 4.
A platelet factor 4 (PF4)-expressing cell line, HELNeo, was derived from the human erythroleukemia cell line, HEL. This was achieved by stable transfection of HEL cells with a construct containing the rat PF4 promoter driving the gene coding for resistance to neomycin, followed by selection of neomycin-resistant clones. HELNeo cells were all nonadhering and about 5% of the cells had polyploid nuclei (> or = 8N), as compared with 1% in HEL cells. Immunohistochemistry showed that about 90% of the HELNeo cells contained PF4, whereas only approximately 5% of the HEL cells contained PF4. No significant parallel enrichment was observed for other megakaryocytic markers, such as the glycoprotein complex IIb/IIIa, von Willebrand factor, and platelet activation-dependent granule to external membrane glycoprotein (PADGEM), which were present to a similar extent in both HEL and HELNeo lines. The increased expression of PF4 in HELNeo cells was confirmed by transient expression assays and was associated with a fivefold increase in trans-acting factors binding to the PF4 promoter. These cells should be a rich source for purifying trans-acting factors binding to the PF4 gene. Moreover, our study shows how a lineage-specific promoter may be used to generate lineage-specific cell lines from a multilineage hematopoietic cell line.
Ravid
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Department of Biology, Massachusetts Institute of Technology, Cambridge 02139.
Kuter
D J
DJ
Beeler
D L
DL
Doi
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T
Rosenberg
R D
RD
D001483:Base Sequence; D002455:Cell Division; D004268:DNA-Binding Proteins / Q000378:metabolism; D015870:Gene Expression; D006801:Humans; D066298:In Vitro Techniques; D004915:Leukemia, Erythroblastic, Acute / Q000378:metabolism*; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000378:metabolism; D009838:Oligodeoxyribonucleotides / Q000737:chemistry; D010978:Platelet Factor 4 / Q000378:metabolism*; D011003:Ploidies; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000235:genetics; D014158:Transcription, Genetic; D014162:Transfection; D014407:Tumor Cells, Cultured
D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D009838:Oligodeoxyribonucleotides; D012333:RNA, Messenger; D010978:Platelet Factor 4