Differentiation-responsive elements in the 5' region of the mouse tissue plasminogen activator gene confer two-stage regulation by retinoic acid and cyclic AMP in teratocarcinoma cells.
F9 cells induced to differentiate with retinoic acid (RA) increase transcription of the tissue plasminogen activator (t-PA) gene. Further treatment of these cells with cyclic AMP (cAMP) results in an additional stimulation of t-PA gene transcription. To investigate the mechanism of this two-stage regulation, 4 kilobase pairs (kbp) of 5'-flanking sequence from the murine t-PA gene was isolated. Two major start sites for transcription were found, neither of which depended on a classical TATA motif for correct initiation. By using transient transfection assays, it was determined that 4-kbp of flanking sequence could confer on reporter genes the same two-stage differentiation-specific expression as was observed for the endogenous t-PA gene. Deletion analyses of this 4-kbp fragment showed that 190 bp of flanking sequence was sufficient to bestow the same degree of two-stage regulation on reporter gene constructs. Within this region of DNA, sequence analysis revealed a possible cAMP regulatory element, a CTF/NF-1 recognition sequence, two potential Sp1 sites, and five potential binding sites for transcription factor AP-2. The deletion experiments, coupled with the positions of these potential cis-acting elements, suggest that multiple transcription factors, including those that bind to cAMP regulatory element, CTF/NF-1, Sp1, and AP-2 sites, may be involved in regulation of the t-PA gene during F9 cell differentiation.
Rickles
R J
RJ
Department of Molecular Pharmacology, State University of New York, Stony Brook 11794.
Darrow
A L
AL
Strickland
S
S
D000818:Animals; D001483:Base Sequence; D002454:Cell Differentiation / Q000187:drug effects*; D002872:Chromosome Deletion; D002874:Chromosome Mapping; D000242:Cyclic AMP / Q000494:pharmacology*; D004273:DNA, Neoplasm / Q000235:genetics; D005786:Gene Expression Regulation; D051379:Mice; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic / Q000187:drug effects; D013724:Teratoma / Q000235:genetics / Q000473:pathology; D010959:Tissue Plasminogen Activator / Q000235:genetics*; D014158:Transcription, Genetic / Q000187:drug effects; D014212:Tretinoin / Q000494:pharmacology*; D014407:Tumor Cells, Cultured / Q000187:drug effects / Q000473:pathology
D004273:DNA, Neoplasm; D014212:Tretinoin; D000242:Cyclic AMP; D010959:Tissue Plasminogen Activator