Macrophage colony-stimulating factor expression in retrovirally transduced cells is dependent upon both the adherence status of the target cells and its 5' flanking untranslated region.
Numerous cell types retrovirally transduced with macrophage colony-stimulating factor (M-CSF) using LXSN-based vectors showed a variable expression of the transgene. Expression of M-CSF correlated with the cells' adherent status. Transduced adherent cells produced the M-CSF, whereas the non-adherent cells synthesized little M-CSF. Studies showed that the 5'-UTR of the M-CSF gene regulated transgenic M-CSF gene expression. Ligation of this 5'-UTR to the enhanced green fluorescent protein gene (EGFP) caused the expression of EGFP to show the same dichotomy as previously seen with the M-CSF. Transgenic M-CSF was expressed within non-adherent cells when the 5'-UTR was removed from the LXSN vector. Quantitative real-time polymerase chain reaction analysis confirmed that lesser production of M-CSF mRNA occurred within the non-adherent cells than in the adherent cells. This difference was eliminated when the 5'-UTR was removed from the retroviral vector. Our work suggests that this 5'-UTR of the M-CSF gene could be an important way to get transgenic expression within adherent cells, but not in non-adherent cells.
Zhang
Jian-Gang
JG
Diagnostic and Molecular Medicine Healthcare Group, Box 113 Veterans Affairs Medical Center, 5901 E. 7th Street, Long Beach, CA 90822, USA. jiangang.zhang@med.va.gov
Dan
Qinghong
Q
Fong
Timothy C
TC
Williams
Christopher C
CC
Avina
Maria D
MD
Tarbiyat-Boldaji
Mehrdokht
M
Khalaghizadeh
Sakineh
S
Irwin
Michael
M
Nguyen
Amy
A
Zhuang
Jing-Li
JL
Hoa
Neil
N
Wepsic
H Terry
HT
Jadus
Martin R
MR
D020121:5' Untranslated Regions / Q000235:genetics*; D000818:Animals; D002417:Cattle; D002448:Cell Adhesion; D002478:Cells, Cultured; D018432:Drug Resistance, Multiple; D005822:Genetic Vectors; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000235:genetics; D049452:Green Fluorescent Proteins / Q000096:biosynthesis / Q000235:genetics; D006801:Humans; D016173:Macrophage Colony-Stimulating Factor / Q000096:biosynthesis* / Q000235:genetics; D051379:Mice; D012333:RNA, Messenger / Q000096:biosynthesis; D051381:Rats; D012190:Retroviridae / Q000235:genetics*; D014161:Transduction, Genetic
D020121:5' Untranslated Regions; D012333:RNA, Messenger; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; D016173:Macrophage Colony-Stimulating Factor; D016178:Granulocyte-Macrophage Colony-Stimulating Factor