Reserve carbohydrates maintain the viability of Saccharomyces cerevisiae cells during chronological aging.

Glycogen and trehalose are well known to participate in many important cell functions, e.g., protection from stress factors, regulation of cell growth and division, spore formation. Since the aging is a complex process involving many aspects of cell metabolism, it was interesting to study the role of glycogen and trehalose in maintenance of viability of aging cells. We have revealed that cell aging is accompanied by an abrupt fall of glycogen and trehalose contents between the second and third weeks of aging. Simultaneously, we observed a decrease in the activity of glycolytic enzymes, phosphofructokinase and hexokinase. At the same time, the viability of aging cells abruptly declined. Although we found neither glycogen nor trehalose in the cells after the third week of aging, they remained viable for some time, apparently due to development of some compensatory metabolic pathways. In spite of this fact, complete death of the cells occurred by the eighth week of experiment, which confirmed irreplaceability of reserve carbohydrates in yeast cell metabolism. Possible reasons of the inability of aging cells to accumulate glycogen and trehalose are discussed in the work.

Preservation of the in vivo state of mitochondrial network for ex vivo physiological study of mitochondria.

Over the course of many years our laboratory has been engaged in the study of physiological functions of mitochondria ex vivo. We showed that the unavoidable destruction of mitochondrial-reticular network during traditional isolation of the mitochondria diminishes the observable ex vivo changes of mitochondrial processes in vivo. Comparing preparations obtained from quiescent and stressed rats, we found that the great difference in size of assemblies of mitochondria preserved in homogenate disappears when it is diluted for the measurement of respiration. This also leads to a decrease in the difference between respiration of mitochondria from quiescent and stressed animals. We developed a new method that provides ex vivo stable preservation of the in vivo network using a cytochemical procedure on glass-adhered lymphocytes in blood smear. We radically changed the incubation medium for the measurement of dehydrogenase activity that excludes an artefact of succinate dehydrogenase hyperactivation ex vivo by non-physiological components of the traditionally used solution. Our method made it possible to observe ex vivo two- to eightfold increase in succinate dehydrogenase activity by adrenaline in vivo, while the activity of alpha-ketoglutarate dehydrogenase changed reciprocally. The data obtained show that the structure changes of the network play an important role in physiological regulation of mitochondrial functions. Thus, it may be possible to correct mitochondrial dysfunctions in the organism by substances supporting the stability of mitochondrial network. The developed method is non-invasive, informative and, therefore, is convenient for clinical investigations, particularly of mitochondrial diseases.

Evolution of glyoxylate cycle enzymes in Metazoa: evidence of multiple horizontal transfer events and pseudogene formation.

BACKGROUND: The glyoxylate cycle is thought to be present in bacteria, protists, plants, fungi, and nematodes, but not in other Metazoa. However, activity of the glyoxylate cycle enzymes, malate synthase (MS) and isocitrate lyase (ICL), in animal tissues has been reported. In order to clarify the status of the MS and ICL genes in animals and get an insight into their evolution, we undertook a comparative-genomic study. RESULTS: Using sequence similarity searches, we identified MS genes in arthropods, echinoderms, and vertebrates, including platypus and opossum, but not in the numerous sequenced genomes of placental mammals. The regions of the placental mammals' genomes expected to code for malate synthase, as determined by comparison of the gene orders in vertebrate genomes, show clear similarity to the opossum MS sequence but contain stop codons, indicating that the MS gene became a pseudogene in placental mammals. By contrast, the ICL gene is undetectable in animals other than the nematodes that possess a bifunctional, fused ICL-MS gene. Examination of phylogenetic trees of MS and ICL suggests multiple horizontal gene transfer events that probably went in both directions between several bacterial and eukaryotic lineages. The strongest evidence was obtained for the acquisition of the bifunctional ICL-MS gene from an as yet unknown bacterial source with the corresponding operonic organization by the common ancestor of the nematodes. CONCLUSION: The distribution of the MS and ICL genes in animals suggests that either they encode alternative enzymes of the glyoxylate cycle that are not orthologous to the known MS and ICL or the animal MS acquired a new function that remains to be characterized. Regardless of the ultimate solution to this conundrum, the genes for the glyoxylate cycle enzymes present a remarkable variety of evolutionary events including unusual horizontal gene transfer from bacteria to animals.