Figure 4.

An O-GlcNAc modified protein is identified as a common regulatory element. (A) The promoters of target co-regulated genes for human, mouse, and rat were analyzed for common regulatory motifs as described in Section “Materials and Methods.” Three genes that were not co-regulated were used as a negative set to avoid identifying non-regulatory motifs. *Denotes no orthologous gene in rat. (B) TOMTOM was used to assign identified regulatory motifs to known transcription factor binding motifs. Regulatory motif 3 matches the Sp1 DNA binding motif with a p-value of 1.3 × 10⁻⁶. (C) Whole cell lysates from insulin responsive and insulin resistant 3T3-F442a adipocytes were subjected to immunoprecipitation with anti-Sp1 or normal rabbit IgG followed by immunoblotting with anti-Sp1 or anti-O-GlcNAc (RL2). A representative immunoblot is shown (right panel). The ratio of O-GlcNAc modified Sp1 to total Sp1 was quantified using densitometry of independent experiments (N = 4).