Lentivirus-mediated bifunctional cell labeling for in vivo melanoma study.
Lentiviral vectors (LVs) are capable of labeling a broad spectrum of cell types, achieving stable expression of transgenes. However, for in vivo studies, the duration of marker gene expression has been highly variable. We have developed a series of LVs harboring different promoters for expressing reporter gene in mouse cells. Long-term culture and colony formation of several LV-labeled mouse melanoma cells showed that promoters derived from mammalian house-keeping genes, especially those encoding RNA polymerase II (Pol2) and ferritin (FerH), provided the highest consistency for reporter expression. For in vivo studies, primary B16BL6 mouse melanoma were infected with LVs whose luciferase-green fluorescence protein fusion gene (Luc/GFP) was driven by either Pol2 or FerH promoters. When transplanted into syngeneic C57BL/6 mice, Luc/GFP-labeled B16BL6 mouse melanoma cells can be monitored by bioluminescence imaging in vivo, and GFP-positive cells can be isolated from the tumors by fluorescence-activated cell sorter. Pol2-Luc/GFP labeling, while lower in activity, was more sustainable than FerH-Luc/GFP labeling in B16BL6 over consecutive passages into mice. We conclude that Pol-2-Luc/GFP labeling allows long-term in vivo monitoring and tumor cell isolation in immunocompetent mouse melanoma models.
Day
Chi-Ping
CP
Laboratory of Cancer Biology and Genetics, National Cancer Institute, National Institutes of Health, Bethesda, MD, USA.
Carter
John
J
Bonomi
Carrie
C
Esposito
Dominic
D
Crise
Bruce
B
Ortiz-Conde
Betty
B
Hollingshead
Melinda
M
Merlino
Glenn
G
D000818:Animals; D004195:Disease Models, Animal; D017930:Genes, Reporter; D005822:Genetic Vectors / Q000235:genetics / Q000378:metabolism; D006801:Humans; D007202:Indicators and Reagents; D016086:Lentivirus / Q000235:genetics / Q000378:metabolism; D008545:Melanoma / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D051379:Mice; D008810:Mice, Inbred C57BL; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D014161:Transduction, Genetic / Q000379:methods; D019076:Transgenes; D014407:Tumor Cells, Cultured
D007202:Indicators and Reagents; D011993:Recombinant Fusion Proteins