FIG. 4.

Sp1 interacts directly with HDAC1. (A) GST–Sp1(622-788) precipitates HDAC1 from extracts prepared from resting and serum-stimulated Swiss 3T3 cells. HDAC1 was detected by Western blot analysis with the anti-HDAC1 antibody. GST and GST-Sp1(1-621) failed to interact with HDAC1. Input extracts (8%) and pull-down assays without extract were included as controls. GST fusion proteins were detected on the same blot with a GST-specific antibody. The GST proteins (without the Sp1 portion) running in the front of the gel are not shown. (B) Sp1-HDAC1 interaction is slightly reduced in S-phase cells. Sp1 was immunoprecipitated (IP) with anti-Sp1 antibody (Pep2) from extracts prepared from G₀- and S-phase Swiss 3T3 cells. Immunoprecipitated Sp1 and coimmunoprecipitated HDAC1 were visualized on the Western blot with the anti-Sp1 antibody and the anti-HDAC1 antibody, respectively. (C) GST-HDAC1 interacts with purified Sp1 (4 FPU; Promega). Sp1 was visualized by Western blot analysis with the anti-Sp1 antibody. GST-E2F4 failed to interact with recombinant Sp1, while E2F1 binds to Sp1, as previously shown (28). Purified Sp1 (2 FPU) was loaded as a control.