FIG. 1.

HMBA stimulates transcription from the HIV-1 promoter. (A) HMBA induces expression of the luciferase reporter gene driven by the HIV-1 LTR. A stable HeLa-based cell line containing an integrated HIV-1 LTR-luciferase reporter construct was treated with 10 mM HMBA for the indicated time periods. The level of induction (n-fold) in luciferase activity compared to that in untreated cells is shown. (B) The HMBA-induced increase in luciferase activity occurs at the mRNA level. Semiquantitative RT-PCR and Northern blot analyses were performed to detect luciferase mRNA from among total RNA isolated at various time points of a continuous HMBA treatment. The levels of the GAPDH mRNA and 7SK snRNA were also analyzed as internal controls. (C) NE from HMBA-treated HeLa cells significantly increases HIV-1 transcription in vitro. Reaction mixtures contained the transcription template HIV+TAR-G400 and NE prepared from cells treated with HMBA for 0 or 3 h. RNA fragments transcribed from a G-less cassette inserted into the template at a position ∼1 kb downstream of the HIV-1 promoter are indicated.