Transcriptional regulation of rat Na+/H+ exchanger isoform-2 (NHE-2) gene by Sp1 transcription factor

The rat Na+/H+ exchanger isoform-2 (NHE-2) gene promoter lacks a TATA box a nd is very GC rich. A minimal promoter extending from bp -36 to +116 direct s high-level expression of NHE-2 in mouse inner medullary collecting duct ( mIMCD-3) cells. Four Sp1 consensus elements were found in this region. The introduction of mutations within these Sp1 consensus elements and DNA footp rinting revealed that only two of them were utilized and are critical for b asal transcriptional activation in mIMCD-3 cells. The use of Sp1, Sp3, and Sp4 antisera in electrophoretic mobility shift assays demonstrated that Sp1 , Sp3, and Sp4 bound to this minimal promoter. We further analyzed the tran scriptional regulation of NHE-2 by members of the Sp1 multigene family. In Drosophila SL2 cells, which lack endogenous Sp1, the minimal promoter canno t drive transcription. Introduction of Sp1 activated transcription over 100 -fold, suggesting that Sp1 is critical for transcriptional regulation. Howe ver, neither Sp3 nor Sp4 was able to activate transcription in these cells. Furthermore, in mIMCD-3 cells, Sp1-mediated transcriptional activation was repressed by expression of Sp3 and Sp4. These data suggest that Sp1 is cri tical for the basal promoter function of rat NHE-2 and that Sp3 and Sp4 may repress transcriptional activation by competing with Sp1 for binding to co re cis-elements.