Tup1 represses transcription mediated by nonclassical activators
particularly efficiently. (A) The reporter used in the
repression assays contains five Gal4 DNA-binding sites upstream of two
lexA sites which are, in turn, positioned 50 bp upstream from the
natural GAL1 TATA box. (B and
C) Cells harboring the reporter gene were cotransformed
with plasmids expressing Gal4-Tup1 and a variety of LexA-fusion
activators. In each case, LexA-fusion activators consisted of
full-length protein with lexA replacing the respective DNA-binding
domain where applicable. Gal4-Tup1 consisted of Gal4 (residues 1–100)
fused to full-length Tup1. Cells were assayed for
GAL1-lacZ expression by β-galactosidase activity
(B). The numbers represent the average of assays on
three independent colonies performed in duplicate. The standard errors
were typically 10–15%. The fold repression, shown in
C, was calculated as the ratio of values obtained with
and without the repressor function of Tup1.