Differentiation of human hematopoietic cells increases expression of a gene transferred by a retroviral vector.
To study expression of a retroviral vector in human hematopoietic lineages, two established human hematopoietic cell lines (HL60 and K562) and a human adherent stromal cell line (KM101) were infected with the vector pZIP-SV(X). Expression of the transferred neomycin resistance gene (neor) of pZIP-SV(X) was evaluated as the ability of the cells to form colonies (greater than 50 cells) in an agar assay in the presence of the neomycin analogue, G418. After infection, all three cell lines produced colonies resistant to G418. The level of neor mRNA in separate colonies was analyzed by Northern blot analysis. The neor gene transferred by the vector pZIP-SV(X) was expressed in both human hematopoietic and stromal cell lines. In addition, primary adherent human stromal cells infected with pZIP-SV(X) grew in the presence of G418. To determine if differentiation of hematopoietic cells affects expression of the retroviral vector, HL60 cells infected with pZIP-SV(X) were induced to differentiate, and the level of neor mRNA measured. The amount of neor mRNA increased when HL60 cells were induced to differentiate along the granulocytic pathway. Conversely, when HL60 cells were induced toward monocytoid differentiation (TPA), the level of neor mRNA did not significantly increase. We conclude that the neor gene transferred by a retroviral vector, pZIP-SV(X), is functionally expressed. In addition, expression of the transferred neor gene is regulated during myeloid differentiation of HL60 cells.
Holland
C A
CA
Department of Radiation Oncology, University of Massachusetts Medical Center, Worcester 01605.
Chada
S
S
Wright
J
J
Whitney
C
C
Harigaya
K
K
Greenberger
J S
JS
Newburger
P E
PE
D002454:Cell Differentiation; D002460:Cell Line; D004351:Drug Resistance / Q000235:genetics; D005786:Gene Expression Regulation; D005814:Genes, Viral; D006412:Hematopoietic Stem Cells / Q000166:cytology*; D006801:Humans; D015151:Immunoblotting; D009355:Neomycin; D012333:RNA, Messenger / Q000378:metabolism; D012190:Retroviridae / Q000235:genetics*; D014162:Transfection
D012333:RNA, Messenger; D009355:Neomycin