SPECIFIC CIS-ACTING SEQUENCE FOR PHO8 EXPRESSION INTERACTS WITH PHO4 PROTEIN, A POSITIVE REGULATORY FACTOR, IN SACCHAROMYCES-CEREVISIAE

被引:34
|
作者
HAYASHI, N [1 ]
OSHIMA, Y [1 ]
机构
[1] OSAKA UNIV,FAC ENGN,DEPT FERMENTAT TECHNOL,2-1 YAMADAOKA,SUITA,OSAKA 565,JAPAN
关键词
D O I
10.1128/MCB.11.2.785
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
The PHO8 gene of Saccharomyces cerevisiae encodes repressible alkaline phosphatase (rALPase; EC 3.1.3.). The rALPase activity of the cells is two to three times higher in medium containing a low concentration of P(i) than in high-P(i) medium due to transcription of PHO8. The P(i) signals are conveyed to PHO8 by binding of PHO4 protein, a positive regulatory factor, to a promoter region of PHO8 (PHO8p) under the influence of the PHO regulatory circuit. Deletion analysis of PHO8p DNA revealed two separate regulatory regions required for derepression of rALPase located at nucleotide positions -704 to -661 (distal region) and -548 to -502 (proximal region) and an inhibitory region located at -421 to -289 relative to the translation initiation codon. Gel retardation experiments showed that a beta-galactosidase-PHO4 fusion protein binds to a 132-bp PHO8p fragment bearing the proximal region but not to a 226-bp PHO8 DNA bearing the distal region. The fusion protein also binds to a synthetic oligonucleotide having the same 12-bp nucleotide sequence as the PHO8p DNA from positions -536 to -525. The 132-bp PHO8p fragment, connected at position -281 of the 5' upstream region of a HIS5'-'lacZ fused gene, could sense P(i) signals in vivo, but a 20-bp synthetic oligonucleotide having the same sequence from -544 to -525 of the PHO8p DNA could not. Linker insertions in the PHO8p DNA indicated that the 5-bp sequence 5'-CACGT-3' from positions -535 to -531 is essential for binding the beta-galactosidase-PHO4 fusion protein and for derepression of rALPase.
引用
收藏
页码:785 / 794
页数:10
相关论文
共 17 条