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Figure 8 | Retrovirology

Figure 8

From: Effect of SWI/SNF chromatin remodeling complex on HIV-1 Tat activated transcription

Figure 8

Effect of SWI/SNF, Tat, or acetylated Tat on restriction enzyme accessibility. A) A diagram of the transcription experiment. Immobilized templates (800 bp from the 5' U3 region and into the Gag region) were assembled into nucleosomes and SWI/SNF, Tat, or acetylated Tat were added to the reaction. In the absence of transcription, nuc-1 blocks restriction enzyme accessibility of Afl II. During transcription and remodeling of the nucleosome, the Afl II site becomes accessible, and the 3' end sequence is released. This fragment can be detected by southern slot blotting [74] using a probe (end labeled oligonucleotide) spanning the U5 region into Gag. B) In vitro transcription using CEM cells (G1/S extract). Similar to Figure 1, CEM cells were treated with hydroxyurea/nocodazole and samples were processed at 9 h post-release. The extracts (EXT, 200 μg) were supplemented with 200 ng of purified SWI/SNF, plus wild-type Tat (500 ng), or acetylated Tat (500 ng). α-amanitin at 0.1 μg/ml was used to detected RNAPII sensitivity. Clades B and E (a generous gift of J. Hiscott) and the TAR mutant LTR (TM26) were used as the DNA templates.

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