Continuous regulation is required to maintain a given cell state(1,2)
or to allow it to change in response to the environment(3,4). Studies
of the mechanisms underlying such regulation have often been hindered
by the inability to control gene expression at will. Among the inducib
le systems available for regulating gene expression in eukaryotes(5-8)
, the tetracycline (tet) regulatable system has distinct advantages(9-
11). It is highly specific, non-toxic and non-eukaryotic, and conseque
ntly does not have pleiotropic effects on host cell genes. Previously
this system also had drawbacks, as it did not extinguish gene expressi
on completely, precluding the study of toxic or growth-inhibitory gene
products. We report here the development of a facile reversible tetra
cycline-inducible retroviral system (designated RetroTet-ART) in which
activators and repressors together are expressed in cells. Gene expre
ssion can now be actively repressed in the absence of tet and induced
in the presence of tet, as we have engineered distinct dimerization do
mains that allow co-expression of homodimeric tet-regulated transactiv
ators and transrepressors in the same cells, without the formation of
non-functional heterodimers. Using this system, we show that growth ar
rest by the cell cycle inhibitor p16 is reversible and dependent on it
s continuous expression.