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Versatile Retrovirus Vector Systems for Regulated Gene Expression In Vitro and In Vivo

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Abstract

Background

Several plasmid DNA-based mammalian expression systems have recently been developed which make it possible to manipulate gene expression via the administration of exogenous agents. In order to extend the application of these systems, we have developed retroviral vectors which allow for the controlled expression of inserted genes both in vitro and in vivo.

Materials and Methods

Two vector strategies which make use of the tetracycline-regulated gene expression system described by Gossen and Bujard were evaluated. In a first strategy, one virus was generated which encoded the tTA or rtTA transactivator gene product, and a second virus was generated in which expression of the gene of interest was dependent upon tetracycline-responsive transcriptional control elements placed either within the viral LTR or within the proviral transcriptional unit. In a second vector strategy, both components of the tet-regulatable system were incorporated into a single proviral genome in such a way that expression of both the transgene and the transactivator gene product were under control of tet-regulatable control elements.

Results

Both vector strategies resulted in the ability to regulate the expression of inserted genes. In one single virus configuration, gene expression could be regulated over 100× and the level of gene expression in the induced state was comparable to or greater than that achieved with standard LTR-based vectors. The use of different deletions in the viral LTR made it possible to generate a number of vectors which provide for a fourfold range of levels of expression of inserted genes in the induced state. Studies in mice with transduced cells demonstrated that gene expression could be induced in vivo by manipulation of tetracycline for at least 48 days.

Conclusions

The availability of highly transmissible, regulatable retroviral vectors should greatly facilitate studies in which it is of interest to manipulate the expression of specific genes in vitro or in vivo.

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Acknowledgments

We thank H. Bujard for providing the plasmids pUHC13-3, pUHD15-l, and pUHG17-l. Bosc23 cells were a gift of W. Pear. The work was supported by NIH grant UO1A126463. D.L. was supported by a grant from the Deutsches Krebsforschungszentrum.

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Author notes

  1. Dirk Lindemann

    Present address: Institute for Virology and Immunology, University of W ürzburg, Versbacher Str. 7, 97222, Würzburg, Germany

  2. Edward Patriquin &  Sandy Feng

    Present address: The Children’s Hospital, Department of Genetics, Harvard Medical School, Howard Hughes Medical Institute, 320 Longwood Ave., Boston, MA, 02115, USA

Authors and Affiliations

  1. Whitehead Institute for Biomedical Research and Department of Biology, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA

    Dirk Lindemann,  Edward Patriquin,  Sandy Feng &  Richard C. Mulligan

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  1. Dirk Lindemann
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Correspondence to Richard C. Mulligan.

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Lindemann, D., Patriquin, E., Feng, S. et al. Versatile Retrovirus Vector Systems for Regulated Gene Expression In Vitro and In Vivo. Mol Med 3, 466–476 (1997). https://doi.org/10.1007/BF03401693

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