The transcriptional activator Ino2p dissociates from the yeast INM1 promoter in induction

Abstract

Mood stabilizers lithium and valproates are widely used in the treatment of bipolar disorder. It has been shown that these drugs can affect the inositol monophosphatase activity and thus the inositol de novo biosynthesis. However, the molecular mechanism of this action has thus far been vague. As such, characterizing the regulation of the gene encoding inositol monophosphatase at the molecular level can help to understand the bipolar disorder. As the model organism, the inositol monophosphatase is encoded by INM1 in Saccharomyces cerevisiae. In this study, we showed, using real-time reverse transcriptase polymerase chain reaction analysis, that INM1 is expressed in the presence of inositol, suggesting that the presence of inositol is required for INM1 transcriptional activation. We also demonstrated, using chromatin immunoprecipitation, that Ino2p is present at the promoter under uninduced conditions. Upon induction, Ino2p dissociates from the INM1 promoter. Furthermore, chromatin remodelers Ino80p and Snf2p are recruited to INM1 promoter upon induction as well as histone acetylases Gcn5p and Esa1p. Altogether, we have provided the evidence which describes how the transcriptional activator and coactivators participate in INM1 activation.

FIG. 1.

INM1 is expressed in the presence of inositol. INM1 and ACT1 mRNA were detected by qRT-PCR with INO1 and ACT1 probes, respectively. Mid-log phase WT (INO2-FLAG) cells or ino2Δ cells were grown 2 h at 30°C in synthetic complete media without uracil (SC-ura) or synthetic complete media (SC) containing 2% glucose (w/v) with 100 μM myo-inositol or without myo-inositol, respectively. qRT-PCR, real-time reverse transcriptase polymerase chain reaction; WT, wild type.

FIG. 2.

Ino2p departs from the INM1 promoter in induction. Real-time PCR of DNA immunoprecipitated with antibodies against Ino2p-FLAG (α-FLAG) at INM1 promoter and INO1 promoter. All experiments were repeated at least three times and, in each experiment, PCR reactions were done in triplicate. The % of input is graphed as mean±standard deviation.

FIG. 3.

Both INO80 and SWI/SNF chromatin remodelers are recruited to the INM1 promoter in induction. Real-time PCR of DNA immunoprecipitated with antibodies against INO80 (α-Arp8p) and SWI/SNF (α-Snf2p) at INM1 promoter. Annotation was as described in the legend of Figure 2.

FIG. 4.

Both Gcn5p and Esa1p histone acetylases are recruited to the INM1 promoter in induction. Real-time PCR of DNA immunoprecipitated with antibodies against Gcn5p (α-Gcn5p) and Esa1p (α-Esa1p) at INM1 promoter. Annotation was as described in the legend of Figure 2.