H-rev107, which belongs to class II tumor suppressor genes, is ubiquitously expressed in normal cells, but is downregulated in many carcinomas and tumor cell lines. Sequence analysis showed that the murine H-rev107 gene is composed of five exons and four introns. Transfections revealed that 7.6 kb of the H-rev107 promoter directed a high level expression of the reporter gene. There were no significant differences in promoter activity when various 5'-deletion promoter constructs from -7.6 kb to - 113 bp were employed. By further deletion and mutation analysis, we found that the region between -83 and -75 containing a GC-box was essential for promoter activity in NIH3T3 or REF52 fibroblasts expressing H-rev107 at moderate to high levels. Gelshifts demonstrated in vitro binding of Sp1 and Sp3 to this GC-box. Cotransfection of Sp1 and Sp3 functionally stimulated promoter activity in SL2 cells. By chromatin immunoprecipitation assays, we observed in vivo binding of Sp1 and Sp3 to the proximal promoter region in NIH3T3 cells and liver, concluding that the transcription of the H-rev107 gene is dependent on Sp1/Sp3-binding to the -83/-75 GC-box. (C) 2002 Elsevier Science (USA). All rights reserved.