Transcriptional control elements of the rat thymidylate synthase promoter:Evolutionary conservation of regulatory features

The sequence elements that are important for the transcription and regulati on of the rat thymidylate synthase (TS) gene were analyzed. The rat TS prom oter lacks a TATA box and directs transcriptional initiation at multiple si tes between 60 and 20 nt upstream of the AUG translational start codon. Pro moter deletion analyses showed that the region between -100 and -42 nt rela tive to the AUG codon was both necessary and sufficient for high level prom oter activity and was designated the essential promoter region. The essenti al region also had bidirectional promoter activity. Site-directed mutagenes is revealed that four elements were especially important for promoter activ ity. These include Ets motifs at -85 and -50, an Sp1 motif at -80, and an L SF motif that overlapped the upstream Ets and Sp1 motifs. Inactivation of E 2F motifs that are upstream and downstream of the essential promoter region had no measurable effect on promoter activity in transient transfection as says. The rat TS promoter region directed S-phase-specific expression of a stably transfected minigene if a spliceable intron was included in the tran scribed region. When the intron was deleted or the E2F motifs were inactiva ted, expression of the minigene changed very little during the G1 to S tran sition. (C) 2000 Academic Press.