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. 2006 Oct;26(19):7068–7076. doi: 10.1128/MCB.00778-06

FIG. 2.

FIG. 2.

HMBA treatment of HeLa cells disrupts the 7SK snRNP and enhances the binding of P-TEFb to the HIV-1 chromatin template. (A) Treatment of HeLa cells with HMBA induces the dissociation of HEXIM1 and 7SK from P-TEFb. F1C2, a HeLa-based cell line stably expressing CDK9-f, was treated with HMBA for the indicated numbers of hours. CDK9-f, CycT1, HEXIM1, and 7SK present in NEs (lanes 1 to 5) and the anti-FLAG immunoprecipitates (αFLAG IP) (lanes 6 to 10) were detected by Western and Northern blotting. The presence of endogenous CDK9 (endo. CDK9) in NEs was revealed by anti-CDK9 Western blotting. (B) HMBA enhances the binding of P-TEFb to the HIV-1 chromatin template. The HeLa-based cell line with the integrated luciferase reporter gene driven by the HIV-1 LTR was treated with HMBA for 0 or 3 h. ChIP with anti-CDK9 antibody was performed. Three regions corresponding to the promoter region, interior region, and 3′ untranslated region (3′ UTR) of the integrated HIV-1 LTR-luciferase gene, as well as an interior region of the endogenous GAPDH gene, were PCR amplified from the precipitated and purified DNA. Numbers in parentheses indicate nucleotides. Amplified signals from 10% of the input chromatin are also shown.