Expression of 5' UTR mutants of Tat RNA in HeLa P4 cells. A. Top panel depicts the DNA constructs used in the experiments. Deletions in the 5' UTR of Tat RNA are indicated. B. Middle panel shows the activity of newly made Tat that activates LacZ transcription from the HIV-1 LTR in HeLa P4 cells. This was monitored by the β-galactosidase activity (see methods) (at least three independent assays were performed). Note that deletion of either the entire 5' UTR (Tatcod) or the sequences encompassing TAR-pA-PBS-DIS strongly impaired Tat expression by the bicistronic RNA. C. Lower panel reports the Renilla luciferase activity (5' cistron) for each DNA construct. Integrity of the recombinant RNAs has been examined by Northern blotting. (not shown).