SP1-MEDIATED TRANSCRIPTIONAL ACTIVATION IS REPRESSED BY SP3

Sp1, Sp3 (SPR-2) and Sp4 (SPR-1) are human sequence-specific DNA bindi ng proteins with very similar structural features. In this report, we have analyzed Sp3 in direct comparison with Sp1. We have raised antibo dies against both Sp1 and Sp3, and show that Sp3 protein, like Sp1 is expressed in various cell lines. Co-transfection experiments in differ ent mammalian cell lines reveal that in contrast to Sp1 and Sp4, Sp3 i s not able to activate several Sp1 responsive promoters. In addition, Sp3 also fails to activate reporter constructs in Drosophila SL2 cells lacking endogenous Sp factors. Instead, We find that Sp3 represses Sp 1-mediated activation in a linear dose-dependent manner. A mutant of S p3 lacking the DNA binding domain does not affect activation by Sp1, s uggesting that the inhibition is most likely due to the competition wi th Sp1 for their common binding sites. To determine if any structurall y similar domain of Sp3 is able to replace partially homologous domain s of Sp1, we have generated chimeric proteins and tested their activat ion characteristics in gene transfer experiments. It appears that neit her the glutamine-rich domains A and B nor the D domain of Sp1 can be replaced by the homologous regions of Sp3. Our results suggest that Sp 3 is an inhibitory member of the Sp family.