Rapamycin treatment results in GATA factor-independent hyperphosphorylation of the proline utilization pathway activator in Saccharomyces cerevisiae.

Treatment of Saccharomyces cerevisiae cells with the immunosuppressive drug rapamycin results in a variety of cellular changes in response to perceived nutrient deprivation. Among other effects, rapamycin treatment results in the nuclear localization of the global nitrogen activators Gln3p and Nil1p/Gat1p, which leads to expression of nitrogen assimilation genes. The proline utilization (Put) pathway genes were shown to be among the genes induced by rapamycin.

Improved anaerobic use of arginine by Saccharomyces cerevisiae.

Anaerobic arginine catabolism in Saccharomyces cerevisiae was genetically modified to allow assimilation of all four rather than just three of the nitrogen atoms in arginine. This was accomplished by bypassing normal formation of proline, an unusable nitrogen source in the absence of oxygen, and causing formation of glutamate instead. A pro3 ure2 strain expressing a PGK1 promoter-driven PUT2 allele encoding Delta(1)-pyrroline-5-carboxylate dehydrogenase lacking a mitochondrial targeting sequence produced significant cytoplasmic activity, accumulated twice as much intracellular glutamate, and produced twice as much cell mass as the parent when grown anaerobically on limiting arginine as sole nitrogen source.

Conformational changes play a role in regulating the activity of the proline utilization pathway-specific regulator in Saccharomyces cerevisiae.

In Saccharomyces cerevisiae, the ability to use proline as a nitrogen source requires the Put3p transcriptional regulator, which turns on the expression of the proline utilization genes, PUT1 and PUT2, in the presence of the inducer proline and in the absence of preferred nitrogen sources. Changes in target gene expression occur through an alteration in activity of the DNA-bound Put3p, a member of the Zn(II)2Cys6 binuclear cluster family of proteins. Here, we report that the 'on' conformation can be mimicked in the absence of proline by the insertion of an epitope tag in several different places in the protein, as well as by specific amino acid changes that suppress a put3 mutation leading to non-inducibility of the pathway.

Cross-pathway regulation in Saccharomyces cerevisiae: activation of the proline utilization pathway by Ga14p in vivo.

The Put3p and Gal4p transcriptional activators are members of a distinct class of fungal regulators called the Cys(6) Zn(II)(2) binuclear cluster family. This family includes over 50 different Saccharomyces cerevisiae proteins that share a similar domain organization. Gal4p activates the genes of the galactose utilization pathway permitting the use of galactose as the sole source of carbon and energy.

The regulator of the yeast proline utilization pathway is differentially phosphorylated in response to the quality of the nitrogen source.

The proline utilization pathway in Saccharomyces cerevisiae is regulated by the Put3p transcriptional activator in response to the presence of the inducer proline and the quality of the nitrogen source in the growth medium. Put3p is constitutively bound to the promoters of its target genes, PUT1 and PUT2, under all conditions studied but activates transcription to the maximum extent only in the absence of rich nitrogen sources and in the presence of proline (i.e.