PHO4 and PHO2 Protein Interact with
Upstream Sequence of PHO81 Gene
XIA Zan-Xian and AO Shi-Zhou
( State Key Laboratory ofmolecular Biology, Shanghai Institute of
Biochemistry,the Chinese Academy of Sciences, Shanghai 200031, China
)
Abstract PHO4 and PHO2 protein were
overexpressed in E. coli and purified respectively. The gel
retardation assays showed that PHO4 and PHO2 protein could bind to –401 to –289
bp sequence of PHO81 gene promoter respectively. The assays of the
mutant PHO2 proteins, where the Ser-230 was changed into Asp (GST-2SD) or Ala
(GST-2SA), showed the binding patterns were the same as the intact PHO2
protein. Footprinting results located the PHO4 binding site at the –354 to –333
bp sequence of PHO81 gene, PHO2 binding site at the –341 to –334
bp. Therefore the region (–401 to –289 bp) contains the upstream activation
sequence (UAS) of PHO81 gene, and PHO4 protein cooperates with PHO2 in
acting on the upstream sequence of PHO81 gene and both of them
regulate the expression of PHO81 gene.
Key Words PHO81; PHO4 and PHO2; retardation
assay; footprinting assay; upstream activation sequence
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