Sp Family Members Stimulate Transcription of the Hex Gene via Interactions with GC Boxes

Abstract

The 5'-flanking region of the mouse Hex gene was examined in order to identify transcription factors regulating its expression in hepatocytes and haematopoietic cells. We have identified two further GC boxes (GC boxes 3 and 4 at nucleotide positions -149 to -140 and -79 to -70, respectively), i. e. in addition to the two previously determined ones (GC boxes 1 and 2 at nucleotide positions -197 to -188 and -176 to -167, respectively). Luciferase reporter assays revealed that all four GC boxes are transcriptionally active in both MH₁, C₁, rat hepatoma and K562 human chronic myelogenous leukemia cells. Electrophoretic mobility shift assays with specific competitors and antibodies showed that members of the Sp family, namely Spl and Sp3, bind to these GC boxes. Overexpression of Spl and Sp3 in Drosophila SL2 cells stimulated transcription of the Hex gene through interactions with GC boxes 1 to 4, Spl being a more potent activator than Sp3. Thus, we conclude that Spl and Sp3 stimulate transcription of the Hex gene in both MH₁C₁, and K562 cells.