Expression and visualization of a human interferon-beta-enhanced green fluorescent protein chimeric molecule in cultured cells.
We have constructed a recombinant cDNA encoding the chimeric protein between human IFN-beta (HuIFN-beta) and enhanced green fluorescent protein (EGFP) to elucidate the intracellular localization of IFN-beta. Transient expression of the chimeric molecule, HuIFN-beta-EGFP, in L cells demonstrated that the chimeric molecule secreted from the cells had an intact biological activity as far as antiviral effect was concerned. Immunostaining of the transfected cells using anti-HuIFN-beta antibody demonstrated that green-fluorescence was co-localized with the IFN signal and its profile was similar to IFN signals in the cells transfected with HuIFN-beta expressing plasmid DNA. These results indicate that the HuIFN-beta-EGFP chimeric gene was expressed as a chimeric protein and the chimera was transported via the regular secretory pathway in the cells. In other cell types, the fluorescence derived from the chimeric protein was also seen on cytoplasmic vesicular structures. These results suggest that HuIFN-beta-EGFP will be a useful tool to investigate the intracellular trafficking processes of HuIFN-beta in a variety of cell types.
Department of Drug Delivery Research, Graduate School of Pharmaceutical Sciences, Kyoto University, Kyoto, Japan.
Masaru
M
Nakamura
Masato
M
Maruyama
Fumiyoshi
F
Yamashita
Yoshinobu
Y
Takakura
Mitsuru
M
Hashida
Yoshihiko
Y
Watanabe
D000818:Animals; D002478:Cells, Cultured; D018076:DNA, Complementary / Q000235:genetics; D049452:Green Fluorescent Proteins / Q000235:genetics*; D006801:Humans; D016899:Interferon-beta / Q000235:genetics*; D051379:Mice; D018613:Microscopy, Confocal; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000235:genetics; D014162:Transfection
D018076:DNA, Complementary; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D016899:Interferon-beta