Transcriptional regulation of the yeast PHO8 promoter in comparison to the coregulated PHO5 promoter

Abstract

Expression of the PHO8 and PHO5 genes that encode a nonspecific alkaline and acid phosphatase, respectively, is regulated in response to the P(i) concentration in the medium by the same transcription factors. Upon induction by phosphate starvation, both promoters undergo characteristic chromatin remodeling, yet the extent of remodeling at the PHO8 promoter is significantly lower than at PHO5. Despite the coordinate regulation of the two promoters, the PHO8 promoter is almost 10 times weaker than PHO5. Here we show that of two Pho4 binding sites that had been previously mapped at the PHO8 promoter in vitro, only the high affinity one, UASp2, is functional in vivo. Activation of the PHO8 promoter is partially Pho2-dependent. However, unlike at PHO5, Pho4 can bind strongly to its binding site in the absence of Pho2 and remodel chromatin in a Pho2-independent manner. Replacement of the inactive UASp1 element by the UASp1 element from the PHO5 promoter results in more extensive chromatin remodeling and a concomitant 2-fold increase in promoter activity. In contrast, replacement of the high affinity UASp2 site with the corresponding site from PHO5 precludes chromatin remodeling completely and as a consequence promoter activation, despite efficient binding of Pho4 to this site. Deletion of the promoter region normally covered by nucleosomes -3 and -2 results in a 2-fold increase in promoter activity, further supporting a repressive role of these nucleosomes. These data show that there can be strong binding of Pho4 to a UAS element without any chromatin remodeling and promoter activation. The close correlation between promoter activity and the extent of chromatin disruption strongly suggests that the low level of PHO8 induction in comparison with PHO5 is partly due to the inability of Pho4 to achieve complete chromatin remodeling at this promoter.