Basidiomycetous and ascomycetous yeast species were tested for manganese tolerance. Basidiomycetous Cryptococcus humicola, Cryptococcus terricola, Cryptococcus curvatus and ascomycetous Candida maltosa, Kluyveromyces marxianus, Kuraishia capsulata, Lindnera fabianii and Sacharomyces cerevisiae were able to grow at manganese excess (2.5 mmol/L), while the growth of basidiomycetous Rhodotorula bogoriensis was completely suppressed. The lag phase duration increased and the exponential growth rate decreased at manganese excess. The increase of cell size and enlargement of vacuoles were characteristics for the cells grown at manganese excess. The alterations in inorganic polyphosphate content and cellular localization were studied. L. fabianii, K. capsulata, C. maltosa, and Cr. humicola accumulated the higher amounts of inorganic polyphosphates, while Cr. terricola and Cr. curvatus demonstrated no such accumulation. The polyphosphate content in the cell wall tested by DAPI staining increased in all species under the study; however, this effect was more pronounced in Cr. terricola and Cr. curvatus. The accumulation of Mg(2+) in the cell wall under Mn(2+) excess was observed in Cr. humicola, Cr. curvatus and Cr. terricola. The accumulation of polyphosphate and magnesium in the cell wall was supposed to be a factor of manganese tolerance in yeasts.

• MeSH
• buněčná stěna chemie   MeSH
• hořčík metabolismus   MeSH
• kvasinky cytologie   účinky léků   růst a vývoj   metabolismus   MeSH
• mangan metabolismus   MeSH
• polyfosfáty metabolismus   MeSH
• tolerance léku * MeSH
• vakuoly metabolismus   ultrastruktura   MeSH
• Publikační typ
• časopisecké články MeSH

The basidiomycetous yeast Cryptococcus humicola was shown to be tolerant to manganese, cobalt, nickel, zinc, lanthanum, and cadmium cations at a concentration of 2.5 mmol/L, which is toxic for many yeasts. The basidiomycetous yeast Cryptococcus terreus was sensitive to all these ions and did not grow at the above concentration. In the presence of heavy metal cations, С. humicola, as opposed to C. terreus, was characterized by the higher content of acid-soluble inorganic polyphosphates. In vivo 4',6'-diamino-2-phenylindole dihydrochloride staining revealed polyphosphate accumulation in the cell wall and cytoplasmic inclusions of С. humicola in the presence of heavy metals. In C. terreus, polyphosphates in the presence of heavy metals accumulate mainly in vacuoles, which results in morphological changes in these organelles and, probably, disturbance of their function. The role of polyphosphate accumulation and cellular localization as factors of heavy metal tolerance of Cryptococcus humicola is discussed.

• MeSH
• antifungální látky metabolismus   toxicita   MeSH
• buněčná stěna chemie   MeSH
• Cryptococcus účinky léků   růst a vývoj   metabolismus   MeSH
• cytoplazma chemie   MeSH
• organely chemie   MeSH
• polyfosfáty metabolismus   MeSH
• těžké kovy metabolismus   toxicita   MeSH
• tolerance léku * MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Inorganic polyphosphate is involved in architecture and functioning of yeast cell wall. The strain of Saccharomyces cerevisiae constitutively overexpressing acid phosphatase Pho5 was constructed for studying the Pho5 properties and its possible participation in polyphosphate metabolism. The parent strain was transformed by the vector carrying the PHO5 gene under a strong constitutive promoter of glyceraldehyde-3-phosphate dehydrogenase of S. cerevisiae. The culture liquid and biomass of transformant strain contained approximately equal total acid phosphatase activity. The levels of acid phosphatase activity associated with the cell wall and culture liquid increased in the transformant strain compared to the parent strain ~ 10- and 20-fold, respectively. The Pho5 preparation (specific activity of 46 U/mg protein and yield of 95 U/L) was obtained from culture liquid of overproducing strain. The overproducing strain had no changes in polyphosphate level. The activity of Pho5 with long-chained polyP was negligible. We concluded that Pho5 is not involved in polyphosphate metabolism. Purified Pho5 showed a similar activity with p-nitrophenylphosphate, ATP, ADP, glycerophosphate, and glucose-6-phosphate. The substrate specificity of Pho5 and its extracellular localization suggest its function: the hydrolysis of organic compounds with phosphoester bonds at phosphate limitation.

• MeSH
• kyselá fosfatasa genetika   izolace a purifikace   metabolismus   MeSH
• polyfosfáty metabolismus   MeSH
• promotorové oblasti (genetika) genetika   MeSH
• rekombinantní fúzní proteiny genetika   izolace a purifikace   metabolismus   MeSH
• Saccharomyces cerevisiae - proteiny genetika   izolace a purifikace   metabolismus   MeSH
• Saccharomyces cerevisiae enzymologie   metabolismus   MeSH
• substrátová specifita MeSH
• Publikační typ
• časopisecké články MeSH

The yeast Saccharomyces cerevisiae accumulates the high levels of inorganic polyphosphates (polyPs) performing in the cells numerous functions, including phosphate and energy storage. The effects of vacuolar membrane ATPase (V-ATPase) dysfunction were studied on polyP accumulation under short-term cultivation in the Pi-excess media after Pi starvation. The addition of bafilomycin A1, a specific inhibitor of V-ATPase, to the medium with glucose resulted in strong inhibition of the synthesis of long-chain polyP and in substantial suppression of short-chain polyP. The addition of bafilomycin to the medium with ethanol resulted in decreased accumulation of high-molecular polyP, while the accumulation of low-molecular polyP was not affected. The levels of polyP synthesis in the mutant strain with a deletion in the vma2 gene encoding a V-ATPase subunit were significantly lower than in the parent strain in the media with glucose and with ethanol. The synthesis of the longest chain polyP was not observed in the mutant cells. The synthesis of only the low-polymer acid-soluble polyP fraction occurred in the cells of the mutant strain. However, the level of polyP1 was nearly tenfold lower than compared to the cells of the parent strain. Both bafilomycin A1 and the mutation in vacuolar ATPase subunit vma2 lead to a considerable decrease of cellular polyP accumulation. Thus, the defects in ΔμH(+) formation on the vacuolar membrane resulted in the decrease of polyP biosynthesis in S. cerevisiae.

• MeSH
• delece genu MeSH
• ethanol metabolismus   MeSH
• inhibitory enzymů metabolismus   MeSH
• kultivační média chemie   MeSH
• makrolidy metabolismus   MeSH
• podjednotky proteinů genetika   metabolismus   MeSH
• polyfosfáty metabolismus   MeSH
• Saccharomyces cerevisiae enzymologie   metabolismus   MeSH
• vakuolární protonové ATPasy genetika   metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Phosphorylation plays a crucial role in the regulation of many fundamental cellular processes. Phosphorylation levels are increased in many cancer cells where they may promote changes in mitochondrial homeostasis. Proteomic studies on various types of cancer identified 17 phosphorylation sites within the human ATP-dependent protease Lon, which degrades misfolded, unassembled and oxidatively damaged proteins in mitochondria. Most of these sites were found in Lon's N-terminal (NTD) and ATPase domains, though little is known about the effects on their function. By combining the biochemical and cryo-electron microscopy studies, we show the effect of Tyr186 and Tyr394 phosphorylations in Lon's NTD, which greatly reduce all Lon activities without affecting its ability to bind substrates or perturbing its tertiary structure. A substantial reduction in Lon's activities is also observed in the presence of polyphosphate, whose amount significantly increases in cancer cells. Our study thus provides an insight into the possible fine-tuning of Lon activities in human diseases, which highlights Lon's importance in maintaining proteostasis in mitochondria.

• MeSH
• elektronová kryomikroskopie MeSH
• fosforylace MeSH
• lidé MeSH
• mitochondrie * metabolismus   MeSH
• polyfosfáty * metabolismus   MeSH
• proteasa La * metabolismus   MeSH
• proteinové domény MeSH
• tyrosin * metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

The green algal genus Cylindrocystis is widespread in various types of environments, including extreme habitats. However, very little is known about its diversity, especially in polar regions. In the present study, we isolated seven new Cylindrocystis-like strains from terrestrial and freshwater habitats in Svalbard (High Arctic). We aimed to compare the new isolates on a molecular (rbcL and 18S rDNA), morphological (light and confocal laser scanning microscopy), and cytological (Raman microscopy) basis. Our results demonstrated that the Arctic Cylindrocystis were not of a monophyletic origin and that the studied strains clustered within two clades (tentatively named the soil and freshwater/glacier clades) and four separate lineages. Morphological data (cell size, shape, and chloroplast morphology) supported the presence of several distinct taxa among the new isolates. Moreover, the results showed that the Arctic Cylindrocystis strains were closely related to strains originating from the temperate zone, indicating high ecological versatility and successful long-distance dispersal of the genus. Large amounts of inorganic polyphosphate (polyP) grains were detected within the chloroplasts of the cultured Arctic Cylindrocystis strains, suggesting effective luxury uptake of phosphorus. Additionally, various intracellular structures were identified using Raman microscopy and cytochemical and fluorescent staining. This study represents the first attempt to combine molecular, morphological, ecological, and biogeographical data for Arctic Cylindrocystis. Our novel cytological observations partially explain the success of Cylindrocystis-like microalgae in polar regions.

• MeSH
• Chlorophyta * MeSH
• fylogeneze MeSH
• polyfosfáty MeSH
• Streptophyta * MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Geografické názvy
• Arktida MeSH
• Svalbard MeSH

Photosynthetic energy conversion and the resulting photoautotrophic growth of green algae can only occur in daylight, but DNA replication, nuclear and cellular divisions occur often during the night. With such a light/dark regime, an algal culture becomes synchronized. In this study, using synchronized cultures of the green alga Desmodesmus quadricauda, the dynamics of starch, lipid, polyphosphate, and guanine pools were investigated during the cell cycle by two independent methodologies; conventional biochemical analyzes of cell suspensions and confocal Raman microscopy of single algal cells. Raman microscopy reports not only on mean concentrations, but also on the distribution of pools within cells. This is more sensitive in detecting lipids than biochemical analysis, but both methods-as well as conventional fluorescence microscopy-were comparable in detecting polyphosphates. Discrepancies in the detection of starch by Raman microscopy are discussed. The power of Raman microscopy was proven to be particularly valuable in the detection of guanine, which was traceable by its unique vibrational signature. Guanine microcrystals occurred specifically at around the time of DNA replication and prior to nuclear division. Interestingly, guanine crystals co-localized with polyphosphates in the vicinity of nuclei around the time of nuclear division.

• MeSH
• buněčná stěna chemie   MeSH
• buněčný cyklus * MeSH
• časové faktory MeSH
• Chlorophyta cytologie   růst a vývoj   MeSH
• guanin analýza   MeSH
• lipidová tělíska metabolismus   MeSH
• lipidy analýza   MeSH
• mikroskopie * MeSH
• polyfosfáty analýza   MeSH
• Ramanova spektroskopie * MeSH
• škrob analýza   MeSH
• velikost buňky MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• srovnávací studie MeSH

• MeSH
• glukokinasa MeSH
• Streptomyces aureofaciens enzymologie   MeSH

The objective of this study was to describe the dependence of textural properties (hardness, cohesiveness, and relative adhesiveness) of processed cheese spreads on the proportion of disodium phosphate (DSP), tetrasodium diphosphate (TSPP), and sodium salts of polyphosphate in ternary mixtures of emulsifying salts. Sodium salts of polyphosphate with different mean lengths (n ≈ 5, 9, 13, 20, and 28) were used. Pentasodium triphosphate (PSTP) was used instead of TSPP in the second part of the study. Products with and without pH adjustment were tested (the target pH value was 5.60-5.80). Textural properties of the processed cheese were observed after 2, 9, and 30 d of storage at 6°C. Hardness of the processed cheese with a low content of polyphosphate increased at a specific DSP:TSPP ratio (~1:1 to 3:4). This trend was the same for all the polyphosphates used; only the absolute values of texture parameters were different. The same trends were observed in the ternary mixtures with PSTP, showing lower final values of hardness compared with samples containing TSPP. Hardness and cohesiveness decreased and relative adhesiveness increased in the samples with increased pH values and vice versa; the main trend remained unchanged.

• MeSH
• difosfáty chemie   MeSH
• emulze MeSH
• fosfáty chemie   MeSH
• koncentrace vodíkových iontů MeSH
• manipulace s potravinami metody   MeSH
• polyfosfáty chemie   MeSH
• skladování potravin MeSH
• soli chemie   MeSH
• sýr analýza   MeSH
• tvrdost MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

• MeSH
• DNA MeSH
• polyfosfáty MeSH
• spektrofotometrie ultrafialová MeSH