FIG. 2.

Specific binding of Sfl1 to UAS2. (A). The AGAA sequences (bp −1106 to −1103) in pLG-UAS2 were replaced with CGCA by site-directed mutagenesis. The resulting construct, pLG-UAS2a, and also the parental pLG-UAS2 construct were transformed into wild type (WT) and sfl1Δ cells. β-Galactosidase activity was determined under repressed conditions by using three independent colonies. (B) ChIP assays for Sfl1. pLG-UAS2 or pLG-UAS2a and plasmid pRS325-SFL1-HA or pRS323-SFL1, expressing Sfl1-HA or Sfl1 from its own promoter, respectively, were cotransformed into wild-type cells. Transformants were grown to mid-log phase in synthetic medium with 2% glucose and were treated with formaldehyde to cross-link DNA and proteins. An α-HA ChIP assay was performed, and the UAS2 region was PCR amplified by using purified DNA to determine Sfll binding to UAS2.