The current study examines the relationship between ERRalpha1 and the Sp1 transcription factor. The transfection of ERRalpha1 enhances Sp1-DNA complex formation as visualized by electrophoresis mobility shift assay (EMSA). In addition, luciferase activity under the control of three consensus sites for Sp1 binding is activated by ERRalpha1 transfection. By looking further upstream from this point we have found that ERRalpha1 stimulates the upregulation of Sp1 protein expression by activating its transcription. Indeed, human Sp1 promoter-dependent luciferase activity is activated by ERRalpha1 transfection and a DNA-protein complex is observed by EMSA using oligonucleotides encompassing the putative ERRalpha1 binding site on the human Sp1 promoter (-1444/-1433: 5'AGGACATGACCT-3'). In addition, activation of Sp1 promoter-dependent luciferase activity depends on the A/B and C domains of ERRalpha1 as shown with truncated ERRalpha1 cDNAs. This report is the first to demonstrate that ERRalpha1 promotes Sp1 activity through upregulation of Sp1 expression at the transcriptional level. (C) 2005 Elsevier Inc. All rights reserved.