Accumulation of dodecyltriphenylphosphonium in mitochondria induces their swelling and ROS-dependent growth inhibition in yeast

Hydrophobic cations with delocalized charge are used to deliver drugs to mitochondria. However, micromolar concentrations of such compounds could be toxic due to their excessive accumulation in mitochondria. We studied possible pathophysiological effects of one such cation, i.e. dodecyltriphenylphosphonium (C₁₂-TPP), in the yeast Saccharomyces cerevisiae. First, we found that C₁₂-TPP induces high-amplitude mitochondrial swelling. The swelling can be prevented by addition of protonophorous uncoupler FCCP or antioxidant alpha-tocopherol, but not other tested antioxidants (N-acetylcysteine and Trolox). Second, FCCP prevents ROS-sensitive fluorescent dye (dichlorofluorescein diacetate) staining of yeast treated with C₁₂-TPP. We also showed that all tested antioxidants partially restore the growth inhibited by C₁₂-TPP. The latter points that ROS rather than the mitochondria swelling limit the growth rate.     

Secondary types of food in the diet of a small-sized mammalian herbivore: from species ecology to habitat inferences

A macroscopic analysis of stomach contents is undertaken in the common vole, Microtus arvalis, obscurus chromosomal form in order to assess the patterns of non-herbivorous foraging activity of a herbivore distributed in different Eurasian biomes. The samples are collected in the Ural Mountains at midsummer (8 localities, 60° 38′ N to 51° 07′ N), and in winter (one locality, 56° 37′ N). Based on the frequency of occurrence and quantitative abundance in stomachs, photoassimilating parts of vascular plants are considered as the primary food category and inflorescences, roots, seeds, mycorrhizal fungi, arthropods, chitin fragments, colorless amorphous matter, and mosses are recognized as the secondary food categories. Although no alternatives to the primary food are found, some of the supplements substitute one another either geographically (fungi in the north versus mature seeds in the south) or by the vertical layers of habitats (seeds and arthropods in the herbaceous layer versus roots and fungi in the shallow layers of soil). Non-staple food consumption in different latitudinal zones at midsummer reflects spatiotemporal dynamics of food availability. From taiga to steppes of the Urals, the common vole is sporadically included in soil foraging but the consumption of mycorrhizal fungi (Glomerales) is only revealed within the zones of dark coniferous forests. The results suggest that the extents of inclusion of a herbivore in soil food webs might vary among ecosystems. In a paleoecological perspective, the analysis of stomach contents in a herbivorous micromammal might reveal fine-scale details of the habitats and provides insights into the stages of the growing season.     

Isolation, phylogeny and evolution of the SymRK gene in the legume genus Lupinus L

SymRK is one of the key genes involved in initial steps of legume symbiotic association with fungi (mycorrhization) and nitrogen-fixing bacteria (nodulation). A large portion of the sequence encoding the extracellular domain of SYMRK was obtained for 38 lupine accessions and 2 outgroups in order to characterize this region, to evaluate its phylogenetic utility, and to examine whether its molecular evolutionary pattern is correlated with rhizobial diversity and specificity in Lupinus. The data suggested that, in Lupinus, SymRK is a single copy gene that shows good phylogenetic potential. Accordingly, SymRK provided additional support to previous molecular phylogenies, and shed additional light on relationships within the Old World group of Lupinus, especially among the African species. Similar to results of other studies, analyses of SymRK sequences were unable to resolve placement of the Florida unifoliolate lineage, whose relationship was weakly supported to either the Old or the New World lupines. Our data are consistent with strong purifying selection operating on SymRK in Lupinus, preserving rather than diversifying its function. Thus, although SymRK was demonstrated to be a vital gene in the early stages of the root-bacterial symbiotic associations, no evidence from present analyses indicate that this gene is involved in changes in rhizobial specificity in Lupinus.     

Calcium signaling in developing embryos: focus on the regulation of cell shape changes and collective movements

During morphogenesis tissues significantly remodel by coordinated cell migrations and cell rearrangements. Central to this problem are cell shape changes that are driven by distinct cytoskeletal reorganization responsible for force generation. Calcium is a versatile and universal messenger that is implicated in the regulation of embryonic development. Although calcium transients accrue clearly and more intensely in tissues undergoing rearrangement/migration, it is far from clear what the role of these calcium signals is. Here we summarize the evidence implicating calcium participation in tissue movements, cell shape changes and the reorganization of contractile cytoskeletal elements in developing embryos. We also discuss a novel hypothesis that short-lived calcium spikes are required in cells and tissues undergoing migration and rearrangements as a fine tuning response mechanism to prevent local, abnormally high fluctuations in cytoskeletal activities.     

Family-wide chemical profiling and structural analysis of PARP and tankyrase inhibitors

Inhibitors of poly-ADP-ribose polymerase (PARP) family proteins are currently in clinical trials as cancer therapeutics, yet the specificity of many of these compounds is unknown. Here we evaluated a series of 185 small-molecule inhibitors, including research reagents and compounds being tested clinically, for the ability to bind to the catalytic domains of 13 of the 17 human PARP family members including the tankyrases, TNKS1 and TNKS2. Many of the best-known inhibitors, including TIQ-A, 6(5H)-phenanthridinone, olaparib, ABT-888 and rucaparib, bound to several PARP family members, suggesting that these molecules lack specificity and have promiscuous inhibitory activity. We also determined X-ray crystal structures for five TNKS2 ligand complexes and four PARP14 ligand complexes. In addition to showing that the majority of PARP inhibitors bind multiple targets, these results provide insight into the design of new inhibitors.     

Mitochondrially-encoded protein Var1 promotes loss of respiratory function in Saccharomyces cerevisiae under stressful conditions

Stressed Saccharomyces cerevisiae cells easily lose respiratory function due to deletions in mitochondrial DNA, and this increases their general stress resistance. Is the loss active? We found that erythromycin (an inhibitor of mitochondrial translation) prevents the loss in control cells but not in the ones expressing mitochondrially-encoded protein Var1 in the nucleus. Var1 is a component of mitochondrial ribosomes; it is hydrophilic, positively charged, and prone to aggregation. Addition of DNase altered Var1 content in a preparation of mitochondrial nucleoids. Our data indicate that Var1 physically interacts with mitochondrial DNA and under stress negatively regulates its maintenance.     

In vivo investigation of homocysteine metabolism to polyamines by high-resolution accurate mass spectrometry and stable isotope labeling

Polyamines are essential polycations, playing important roles in mammalian physiology. Theoretically, the involvement of homocysteine in polyamine synthesis via S-adenosylmethionine is possible; however, to our knowledge, it has not been established experimentally. Here, we propose an original approach for investigation of homocysteine metabolites in an animal model. The method is based on the combination of isotope-labeled homocysteine supplementation and high-resolution accurate mass spectrometry analysis. Structural identity of the isotope-labeled metabolites was confirmed by accurate mass measurements of molecular and fragment ions and comparison of the retention times and tandem mass spectrometry fragmentation patterns. Isotope-labeled methionine, spermidine, and spermine were detected in all investigated plasma and tissue samples. The induction of moderate hyperhomocysteinemia leads to an alteration in polyamine levels in a different manner. The involvement of homocysteine in polyamine synthesis and modulation of polyamine levels could contribute to a better understanding of the mechanisms connected with homocysteine toxicity.     

Erythrocyte concentrates, low in buffy coat: production, preservation and evaluation of its quality

Buffy coat-poor packed red cells were prepared from fresh ACD-, ACD-AG- and EDTA-blood, than resuspended with a preservation solution, containing glucose, adenine, guanosine, sucrose, citric acid and sodium citrate and stored at 4 degrees C for 6 weeks. The survival rate of resuspended red cells from ACD-AG-blood amounted to 77% after 6 weeks of storage. The ATP content of resuspended red cells was approximately 25% lower than in ACD-AG whole blood during storage caused probably by increased ATP consuming reactions at the red cell membrane. The P2G-content of resuspended red cells from ACD- and ACD-AG-blood decreased above 50% of the normal level during the first week, as fast as in ACD- and ACD-AG whole blood. The P2G-breakdown in red cells from EDTA-blood was delayed for a week due to the higher pH as in CPD blood. Additions of xylitol, inorganic phosphate, and bicarbonate in 6, 5 and 20 mM final concentrations in the red cell suspensions and an increased pH at the same time delayed the breakdown of ATP and P2G. Packed red cells can be administered fast enough at hematocrits to 0.60 that will be achieved by adding 50 to 100 ml preservation solution. Leukocytes and thrombocytes were reduceds to 70 to 80%. With increasing rate of reduction a higher loss of red cells occured. Buffy coat-poor red cell concentrate contains only few microaggregates. It diminishes the risc of febrile transfusion reactions and delays the appearance of alloimmunisation. The circulatory overload of patients is less frequent than after transfusions of red cell resuspensions containing a large resuspension volume.     

Effect of Gamma Irradiation on the Preference Behavior and Lipid Metabolism in Grey Garden Slug Deroceras reticulatum Mull.

We studied the preference behavior of slugs Deroceras reticulatum in experimental light, thermal, and salt gradients. The zone of each gradients preferred by the majority of individuals had the lowest contents of total lipids and total fatty acids in the tissues as compared to animals in any other zones. Hard gamma irradiation of slugs D. reticulatum proved to induce qualitative changes in salt preference, to decrease photoresponse in light gradients, and to increase lipid content in tissues.     

Effects of cold exposure in vivo and uncouplers and recouplers in vitro on potato tuber mitochondria

Effects of cold exposure in vivo and treatment with laurate, carboxyatractylate, atractylate, nucleotides, and BSA in vitro on potato tuber mitochondria have been studied. Cold exposure of tubers for 48-96 h resulted in some uncoupling that could be reversed completely by BSA and partially by ADP, ATP, UDP, carboxyatractylate, and atractylate. UDP was less effective than ADP and ATP, and atractylate was less effective than carboxyatractylate. The recoupling effects of nucleotides were absent when the nucleotides were added after carboxyatractylate. GDP, UDP, and CDP did not recouple mitochondria from either the control or the cold-exposed tubers. This indicates that the cold-induced fatty acid-mediated uncoupling in potato tuber mitochondria is partially due to the operation of the ATP/ADP antiporter. As to the plant uncoupling protein, its contribution to the uncoupling in tuber is negligible or, under the conditions used, somehow desensitized to nucleotides.