Versatile retrovirus vector systems for regulated gene expression in vitro and in vivo

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.

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.

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 100X 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 four-fold 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.

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.