Regulation of the Cln3-Cdc28 kinase by cAMP in Saccharomyces cerevisiae.

The yeast Saccharomyces cerevisiae grows at widely varying rates in different growth media. In order to maintain a relatively constant cell size, yeast cells must regulate the rate of progress through the cell cycle to match changes in growth rate, moving quickly through G1 in rich medium, and slowly in poor medium. We have examined connections between nutrients, and the expression and activity of Cln3-Cdc28 kinase that regulates the G1-S boundary of the cell cycle in yeast, a point referred to as Start. We find that Cln3 protein levels are highest in glucose and lower in poorer carbon sources. This regulation involves both transcriptional and post-transcriptional control. Although the Ras-cAMP pathway does not appear to affect CLN3 transcription, cAMP increases Cln3 protein levels and Cln3-Cdc28 kinase activity. This regulation requires untranslated regions of the CLN3 message, and can be explained by changes in protein synthesis rates caused by cAMP. A model for CLN3 regulation and function is presented in which CLN3 regulates G1 length in response to nutrients.

Transcriptional regulation of CLN3 expression by glucose in Saccharomyces cerevisiae.

In Saccharomyces cerevisiae, the transition from the G1 phase of the mitotic cycle into S phase is controlled by a set of G1 cyclins that regulate the activity of the protein kinase encoded by CDC28. Yeast cells regulate progress through the G1/S boundary in response to nutrients, moving quickly through G1 in glucose medium and more slowly in poorer medium. We have examined connections between glucose and the level of the message encoding Cln3, a G1 cyclin. We found that glucose positively regulates CLN3 mRNA levels through a set of repeated AAGAAAAA (A2GA5) elements within the CLN3 promoter. Mutations in these sequences reduce both transcriptional activation and specific interaction between CLN3 promoter elements and proteins in yeast extracts. Creation of five point mutations, replacing the G's within these repeats with T's, in the CLN3 promoter substantially reduces CLN3 expression in glucose medium and inhibits the ability of the cells to maintain a constant size when shifted into glucose.

Activation of the Ras/cyclic AMP pathway in the yeast Saccharomyces cerevisiae does not prevent G1 arrest in response to nitrogen starvation.

Cells carrying mutations that activate the Ras/cyclic AMP (Ras/cAMP) pathway fail to accumulate in G1 as unbudded cells and lose viability in response to nitrogen starvation. This observation has led to the idea that cells carrying this type of mutation are sensitive to nitrogen starvation because they are unable to appropriately arrest in G1. In this study, we tested predictions made by this model. We found that cells with activating Ras/cAMP pathway mutations do not continue to divide after nitrogen starvation, show a normal decrease in steady state levels of START-specific transcripts, and are not rescued by removal of cAMP during nitrogen starvation. These findings are inconsistent with the idea that activation of the Ras/cAMP pathway prevents growth arrest in cells starved for nitrogen. Our finding that cells with an active Ras/cAMP pathway have dramatically reduced amino acid stores suggests an alternative model. We propose that cells at high cAMP levels are unable to store sufficient nutrients to allow return to the G1 phase of the cell cycle when they are suddenly deprived of nitrogen. It is this inability to return to G1, rather than a failure to arrest, which leaves cells at different points in the cell cycle following nitrogen starvation.

Changes in gene expression in the Ras/adenylate cyclase system of Saccharomyces cerevisiae: correlation with cAMP levels and growth arrest.

Levels of cyclic 3',5'-cyclic monophosphate (cAMP) play an important role in the decision to enter the mitotic cycle in the yeast, Saccharomyces cerevisiae. In addition to growth arrest at stationary phase, S. cerevisiae transiently arrest growth as they shift from fermentative to oxidative metabolism (the diauxic shift). Experiments examining the role of cAMP in growth arrest at the diauxic shift show the following: 1) yeast lower cAMP levels as they exhaust their glucose supply and shift to oxidative metabolism of ethanol, 2) a reduction in cAMP is essential for traversing the diauxic shift, 3) the decrease in adenylate cyclase activity is associated with a decrease in the expression of CYR1 and CDC25, two positive regulators of cAMP levels and an increase in the expression of IRA1 and IRA2, two negative regulators of intracellular cAMP, 4) mutants carrying disruptions in IRA1 and IRA2 were unable to arrest cell division at the diauxic shift and were unable to progress into the oxidative phase of growth. These results indicate that changes cAMP levels are important in regulation of growth arrest at the diauxic shift and that changes in gene expression plays a role in the regulation of the Ras/adenylate cyclase system.

[Problems of instituting insulin therapy in advanced age]. / Probleme bei der Realisierung einer Insulintherapie im höheren Lebensalter.

The population of diabetics of a district was analysed concerning the age structure, the proportion of the insulin-dependent patients as well as the insulin-dependent diabetics older than 60 years were analysed with regard to the degree of the realization of insulinisation. 13.8% of all diabetics with an age older than 60 years are insulin-dependent. In 71.5% of them the insulinisation is realized. Only in 43% of all insulin-dependent older diabetics an autoinjection of insulin is possible. The results are discussed and propositions for changing the situation are submitted.