Involvement of the TOR signaling pathway in metabolism regulation of Saccharomyces cerevisiae grown under carbohydrate-limited conditions

The lack of nutrients and energy in the cell is tightly connected with periods of their low or high levels. This forces the switch from anabolism to catabolism and vise versa. One important protein that regulates this switch is a protein kinase TOR (target of rapamycin). TOR signaling pathway is a highly conservative controller of many functions, including the intensity of metabolism and stress resistance in a variety of organisms from yeast to humans. It is known that atypical kinase TOR is a component of a complex signaling system, which normally regulates growth and proliferation of cells. Identification of TOR, as an integral component of РІЗК/АКТ pathway and existence of cross anticancer action between p53 and TOR signaling pathways demonstrate the unique role of TOR kinase during cell growth. In fact, various aspects of TOR kinase regulation are examined. As an example, TOR kinase interaction with the basic cellular signaling cascades makes it a useful target for treatment of cancer, diabetes and obesity. It is known that TOR is a nutrient sensor in the cell, which participates in the regulation of signal transduction in response to nutrients (e.g. proteins and amino acids). However, interplay between substances like carbohydrates, that are a major source of energy and carbon for cell, and TOR-signaling pathway remains poorly studied. Metabolic activity is an indicator of intensity of redox processes in the cell. Since the intracellular redox balance depends on the total metabolic activity of cells, we have studied the metabolic activity in the presence of low concentrations of carbohydrates. In the presence of 0.1% carbohydrates in the yeast cultivation medium, fructose-grown parental strain and single mutants demonstrated metabolic activities lower than respective glucose-grown. It can serve as a confirmation of our assumptions about pre-adaptations of fructose-grown yeast cells to carbonyl/oxidative stress. Also, in most cases this parameter was higher in mutant cells compared with the parental strain. Glucose-6-phosphate dehydrogenase is a key enzyme of pentose phosphate pathway and plays an important role in maintaining proper intracellular pool of reduced coenzyme NADPH. Changes in the enzyme activity can be used as a potential biomarker of carbonyl/oxidative stress. It should also be noted that in most cases cells grown in the medium with fructose had lower activity of the enzyme than those in the presence of glucose. Interestingly, the activity of glucose-6-phosphate in mutant strains was lower compared to the parental strain. That can be explained by the redirection of carbohydrates towards non-enzymatic transformations. Thus, under starvation conditions with fructose all studied strains grew faster than those in the presence of glucose. It can be suggested that fructose as compared to glucose accelerates aging by higher metabolism intensity, generation of reactive oxygen and carbonyl species. The above suggestion is in a good agreement with the fact that the activity of G6PDH was lower in the presence of fructose, resulting in acceleration of non-enzymatic reactions. Calorie restriction (0.1% carbohydrates in the medium) inhibits TOR signaling pathway. Metabolic activity is higher, and the activity of glucose-6-phosphate dehydrogenase is lower in the mutant cells compared with the parental strain. This suggests activating of compensatory mechanisms, including protein Snf1p/AMP, Sch9, PKA, MAP that in some way compensates the lack of TOR signaling pathway.