Stable and strictly controlled expression of LTR-flanked autoregulated expression cassettes upon adenoviral transfer.
An autoregulatory bidirectional expression cassette encoding all components necessary for regulated gene expression in a one-step gene transfer was evaluated for use in adenoviral vectors. Adenoviral vectors transducing this cassette provide about 1000-fold regulation. Regulation could be further improved by integrating the cassette as a retroviral vector into the adenoviral backbone. Moreover, with these adeno/retroviral hybrid vectors, the frequency of chromosomal integration is enhanced and about 1% of infected cells show stable chromosomal integration of the autoregulated cassette. In these stably transduced cells high regulation capacity is maintained. To elucidate the molecular mechanism underlying this unexpected observation we investigated the regulation capacity of these cassettes in a viral and non-viral vector background after stable integration into the host's DNA. While naked cassettes show regulated expression that is strongly influenced by the chromosomal surrounding sequences the regulatory capacity of LTR flanked cassettes is highly comparable amongst different cell clones. This strict regulation with little influence from the flanking sequences is obtained when LTR-flanked cassettes are transduced as DNA, by retroviral or by adenoviral infection.
Unsinger
Jacqueline
J
Department of Gene Regulation and Differentiation, GBF-National Research Institute for Biotechnology, Mascheroder Weg 1, 38124 Braunschweig, FRG, Germany.
Lindenmaier
Werner
W
May
Tobias
T
Hauser
Hansjörg
H
Wirth
Dagmar
D
D000256:Adenoviridae / Q000235:genetics* / Q000378:metabolism; D000818:Animals; D005786:Gene Expression Regulation; D018014:Gene Transfer Techniques; D017930:Genes, Reporter; D005822:Genetic Vectors / Q000235:genetics* / Q000378:metabolism; D051379:Mice; D041681:NIH 3T3 Cells; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012190:Retroviridae / Q000235:genetics / Q000378:metabolism; D020079:Terminal Repeat Sequences; D014161:Transduction, Genetic; D019076:Transgenes
D011993:Recombinant Fusion Proteins