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Two upstream activation sites (UASs), UASI and UAS2, were required for full transcriptional activation of STAI in Saccharomyces cerevisiae var. diastaticus. UAS1 was divided into UASI-1 and UASI-2 and conferred the negative regulations of the CYCI promoter [Ahn, J. H., Park, S. H., and Kang, H. S. (1994) MoL Gen. Genet. in press]. In this study, the structural characterization of UASI-1 and the identification of UASl-1 binding factors were carried out. Through gel retardation and DNase I footprinting, three regions (box A, B, and C) bound by cellular factors were identified. When linker insertional mutagenesis was employed on UAS1-1, the region containing box A and B and the structural context were found to be required for UAS function. A specific DNA-protein complex was formed to a synthetic 31-bp DNA fragment containing box A and B. Specific UV-crossfinking confirmed its specific binding and revealed that its approximate molecular mass was 50 kDa.
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