Regulation of the yeast INO1 gene. The products of the INO2, INO4 and OPI1 regulatory genes are not required for repression in response to inositol

The ino2Delta, ino4Delta, opi1Delta, and sin3Delta mutations all affect expression of INO1, a structural gene for inositol-1-phosphate synthase. These same mutations affect other genes of phospholipid biosynthesis that, like INO1, contain the repeated element UAS(INO) (consensus 5' CATGTGAAAT 3'). In this study, we evaluated the effects of these four mutations, singly and in all possible combinations, on growth and expression of INO1. All strains carrying an ino2Delta or ino4Delta mutation, or both, failed to grow in medium lacking inositol. However, when grown in liquid culture in medium containing limiting amounts of inositol, the opi1Delta ino4Delta strain exhibited a level of INO1 expression comparable to, or higher than, the wild-type strain growing under the same conditions. Furthermore, INO1 expression in the opi1Delta ino4Delta strain was repressed in cells grown in medium fully supplemented with both inositol and choline. Similar results were obtained using the opi1Delta ino2Delta ino4Delta strain. Regulation of INO1 was also observed in the absence of the SIN3 gene product. Therefore, while Opi1p, Sin3p, and the Ino2p/Ino4p complex all affect the overall level of INO1 expression in an antagonistic manner, they do not appear to be responsible for transmitting the signal that leads to repression of INO1 in response to inositol. Various models for Opi1p function were tested and no evidence for binding of Opi1p to UAS(INO), or to Ino2p or Ino4p, was obtained.     

Regulation of the cell integrity pathway by rapamycin-sensitive TOR function in budding yeast

The TOR (target of rapamycin) pathway controls cell growth in response to nutrient availability in eukaryotic cells. Inactivation of TOR function by rapamycin or nutrient exhaustion is accompanied by triggering various cellular mechanisms aimed at overcoming the nutrient stress. Here we report that in Saccharomyces cerevisiae the protein kinase C (PKC)-mediated mitogen-activated protein kinase pathway is regulated by TOR function because upon specific Tor1 and Tor2 inhibition by rapamycin, Mpk1 is activated rapidly in a process mediated by Sit4 and Tap42. Osmotic stabilization of the plasma membrane prevents both Mpk1 activation by rapamycin and the growth defect that occurs upon the simultaneous absence of Tor1 and Mpk1 function, suggesting that, at least partially, TOR inhibition is sensed by the PKC pathway at the cell envelope. This process involves activation of cell surface sensors, Rom2, and downstream elements of the mitogen-activated protein kinase cascade. Rapamycin also induces depolarization of the actin cytoskeleton through the TOR proteins, Sit4 and Tap42, in an osmotically suppressible manner. Finally, we show that entry into stationary phase, a physiological situation of nutrient depletion, also leads to the activation of the PKC pathway, and we provide further evidence demonstrating that Mpk1 is essential for viability once cells enter G(0).