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. 2008 Sep 8;105(37):14023–14027. doi: 10.1073/pnas.0806726105

Fig. 3.

Fig. 3.

Nuclease digestions released acetylated FOXP3 from chromatin. (A) Ten million serum-starved transfected or control cells were stimulated with a combination of 50 ng/ml PMA and 1 μM ionomycin or 1 mM 8-bromo-cAMP alone for the indicated time. Isolated chromatin fractions were digested with 20 units of DNase I. The released chromatin-bound proteins in solution were separated by SDS/PAGE, transferred, and immunobloted with anti-HA-HRP conjugated antibody. (B) MNase-mediated release of acetylated FOXP3 from chromatin. Ten million HA-FOXP3a- and -3b-transfected or untransfected Jurkat T cells were stimulated as indicated. The chromatin fractions were digested with 10 units of MNase (Roche), the released chromatin digested material was tested for FOXP3 by immunoblotting with anti-HA-HRP antibody, and its acetylation was detected by immunoblotting with anti-acetyl lysine-specific mAb-Ac-K-103 (Upstate Biotechnology).