Construction and characterization of a recombinant adenovirus vector carrying the human preproinsulin gene under the control of the metallothionein gene promoter.
A new adenovirus vector carrying human-preproinsulin (h-PPI) genomic DNA, which was placed under the control of the mouse metallothionein gene promoter, was constructed. In the recombinant virus-infected cells, h-PPI gene expression increased as a function of ZnSO4 concentration. Reversed-phase high-performance liquid chromatography analysis revealed that the recombinant adenovirus-infected cells secreted immature insulin containing proinsulin and incorrectly processed insulin. Tyrosyl phosphorylation of human insulin receptor substrate 1 occurred when HepG2 cells were treated with the cultured medium, indicating that the h-PPI gene product was functionally active in vitro. We also examined the biological activity of the product using diabetic severe combined immunodeficient mice and confirmed that the h-PPI gene product reduced the blood glucose concentration in vivo. This study suggests that the adenovirus vector can be used to express a foreign gene under the control of an external promoter in various human cells.
Yajima
H
H
Faculty of Bioscience and Biotechnology, Tokyo Institute of Technology, 4259, Nagatsuta-cho, Midori-ku, Yokohama, 226, Japan. hhanda@bio.titech.ac.jp
Kosukegawa
A
A
Hoque
M M
MM
Shimojima
T
T
Ishizu
K
K
Takayama
M
M
Sasaki
Y
Y
Sakai
H
H
Otsuka
M
M
Inokuchi
S
S
Handa
H
H
D000256:Adenoviridae; D000818:Animals; D002460:Cell Line; D004274:DNA, Recombinant; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D006801:Humans; D007328:Insulin; D008668:Metallothionein / Q000235:genetics*; D051379:Mice; D011384:Proinsulin / Q000235:genetics*; D011401:Promoter Regions, Genetic / Q000235:genetics; D011498:Protein Precursors / Q000235:genetics*
D004274:DNA, Recombinant; D007328:Insulin; D011498:Protein Precursors; C013737:preproinsulin; D011384:Proinsulin; D008668:Metallothionein