Sp1 and Sp3 transactivate the RET proto-oncogene promoter.
The RET proto-oncogene plays an important role in the initiation and progression of tumors derived from the neural crest. The cis-regulatory elements responsible for RET basal promoter activity have not been identified. To characterize these elements, a RET promoter DNA fragment (-453 to +227bp) was fused to a luciferase reporter and introduced into TT, a neural crest-derived cell line. Sequential 5' deletions of the promoter revealed that optimal expression of the RET promoter in TT cells required only 70bp of sequence upstream of the transcription start site, and contains two Sp1 binding sites. DNase I footprinting, electrophoretic mobility shift analysis (EMSA), and supershift assays revealed that this region binds both Sp1 and its related protein, Sp3. Additionally, RET basal promoter activity was abrogated by removal of these Sp1/Sp3 binding sites. The proximal two GC boxes were sufficient to allow transactivation of the RET promoter in Drosophila SL2 cells. Sp3 expression in these cells caused an additional activation of the promoter. These results demonstrate that the transactivation of the RET promoter within a neural crest-derived cell line is dependent on Sp1 and Sp3.
Andrew
S D
SD
Department of Cancer Genetics, Kolling Institute of Medical Research, Royal North Shore Hospital, NSW 2065, St. Leonards, Australia.
Delhanty
P J
PJ
Mulligan
L M
LM
Robinson
B G
BG
D000818:Animals; D001665:Binding Sites / Q000235:genetics; D002460:Cell Line; D004247:DNA / Q000235:genetics / Q000378:metabolism; D018983:DNA Footprinting; D004268:DNA-Binding Proteins / Q000378:metabolism*; D003851:Deoxyribonucleases / Q000378:metabolism; D029721:Drosophila Proteins; D004591:Electrophoresis, Polyacrylamide Gel; D005786:Gene Expression Regulation; D008156:Luciferases / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D009154:Mutation; D011401:Promoter Regions, Genetic / Q000235:genetics*; D011485:Protein Binding; D011518:Proto-Oncogene Proteins / Q000235:genetics*; D051096:Proto-Oncogene Proteins c-ret; D012313:RNA / Q000235:genetics / Q000378:metabolism; D020794:Receptor Protein-Tyrosine Kinases / Q000235:genetics*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012045:Regulatory Sequences, Nucleic Acid; D012260:Ribonucleases / Q000378:metabolism; D017384:Sequence Deletion; D016329:Sp1 Transcription Factor / Q000378:metabolism*; D051705:Sp3 Transcription Factor; D014157:Transcription Factors / Q000378:metabolism*; D014158:Transcription, Genetic; D015533:Transcriptional Activation
D004268:DNA-Binding Proteins; D029721:Drosophila Proteins; D011518:Proto-Oncogene Proteins; D011993:Recombinant Fusion Proteins; D016329:Sp1 Transcription Factor; D014157:Transcription Factors; D051705:Sp3 Transcription Factor; D012313:RNA; D004247:DNA; D008156:Luciferases; D051096:Proto-Oncogene Proteins c-ret; D020794:Receptor Protein-Tyrosine Kinases; C416649:Ret protein, Drosophila; D003851:Deoxyribonucleases; D012260:Ribonucleases