Screening assay for inhibitors and activators of rna and dna-dependent

Abstract

The invention provides methods for the identification and discovery of agents which are inhibitors and activators of RNA and DNA-dependent nucleic acid polymerases. The essential feature of the invention is the incorporation of a functional polymerase binding site sequence (PBS) into a nucleic acid molecule which is chosen for its ability to confer a discernible characteristic via its sequence specific activity such that the incorporation of the PBS renders the nucleic acid molecule a functional template for a predetermined RNA or DNA-template directed nucleic acid polymerase (1). In the presence of the polymerase, suitable primer molecules, and any necessary accessory molecules, catalytic extension of the strand of nucleic acids complementary to the template occurs, resulting in a partial or total elimination of (or increase in) the characteristic conferring activity of the reporter-template molecule described due to the antisense effects of the complementary strand or other polymerase-mediated effects. Candidate inhibitors and activators are evaluated for their specific effects on polymerase function, versus polymerase-unrelated effects on reporter-template function, by comparing their ability to decrease or increase the extent of the characteristic conferred by the activity of the reporter-template molecule in the assay to their ability to do so in the control situation in which the activity of the polymerase has been eliminated from the assay. A novel method for controlling gene expression and, in general, the activity of a nucleic acid molecule is also disclosed.