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
(1R,2R-diaminocyclohexane)(dihydropyrophosphato) platinum(II), also abbreviated as RRD2, belongs to a class of potent antitumor platinum cytostatics called phosphaplatins. Curiously, several published studies have suggested significant mechanistic differences between phosphaplatins and conventional platinum antitumor drugs. Controversial findings have been published regarding the role of RRD2 binding to DNA in the mechanism of its antiproliferative activity in cancer cells. This prompted us to perform detailed studies to confirm or rule out the role of RRD2 binding to DNA in its antiproliferative effect in cancer cells. Here, we show that RRD2 exhibits excellent antiproliferative activity in various cancer cell lines, with IC50 values in the low micromolar or submicromolar range. Moreover, the results of this study demonstrate that DNA lesions caused by RRD2 contribute to killing cancer cells treated with this phosphaplatin derivative. Additionally, our data indicate that RRD2 accumulates in cancer cells but to a lesser extent than cisplatin. On the other hand, the efficiency of cisplatin and RRD2, after they accumulate in cancer cells, in binding to nuclear DNA is similar. Our results also show that RRD2 in the medium, in which the cells were cultured before RRD2 accumulated inside the cells, remained intact. This result is consistent with the view that RRD2 is activated by releasing free pyrophosphate only in the environment of cancer cells, thereby allowing RRD2 to bind to nuclear DNA.
- MeSH
- cisplatina farmakologie MeSH
- difosfáty farmakologie MeSH
- DNA metabolismus MeSH
- nádory * MeSH
- organoplatinové sloučeniny farmakologie metabolismus MeSH
- oxaliplatin farmakologie MeSH
- platina farmakologie MeSH
- protinádorové látky * farmakologie metabolismus MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Pho91 is a vacuolar phosphate transporter that exports phosphate from the vacuolar lumen to the cytosol in yeast cells. In this study, we have demonstrated the pleiotropic effects of the PHO91 gene knockout in the methylotrophic yeast Ogataea parapolymorpha (Hansenula polymorpha, Ogataea angusta). The content of both acid-soluble and acid-insoluble inorganic polyphosphate (polyP) in the ∆pho91 cells was slightly higher compared to the strain with wild-type PHO91, when the cells were cultivated on glucose. The pho91-Δ mutations both in O. parapolymorpha and in Saccharomyces cerevisiae diminished resistance to cadmium and increased resistance to manganese and peroxide stresses. The cells of the mutant strain of O. parapolymorpha were unable to consume methanol due to the lack of methanol oxidase activity. We speculate that these effects are associated with the inability of mutant cells to mobilize phosphate from the vacuolar pool and/or defects in the signaling pathways involving phosphate, polyP, and inositol polyphosphates.
- MeSH
- genový knockout MeSH
- polyfosfáty * MeSH
- Publikační typ
- časopisecké články MeSH
Podle definice života – množivost (replikace), metabolismus a energetická potřeba a schopnost reagovat na adekvátní podněty mají základní stavební molekuly života společnou hlavní nosnou strukturu: purinovou bázi – adenin, ribózu a dvě fosfátové skupiny. V tomto smyslu může molekula ATP představovat nosič a donor energie a organizátor molekulární výstavby, obnovy a replikace i signální molekulu v mechanismech reaktivity.
According to the definition of life – reproduction (replication), metabolism and energy requirements and the ability to respond to adequate stimuli, the basic building molecules of life have a common principal structure: the purine base - adenine, ribose and two phosphate groups. In this sense, the ATP molecule can represent a carrier and donor of energy and an organizer of molecular construction, renewal and replication, as well as a signalling molecule in reactivity mechanisms.
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
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
The moving volutin (polyphosphate) granules known as "dancing bodies" can be observed in the vacuoles of the yeast cells. The aim of work was to study the effects of cultivation conditions and influences of physico-chemical factors on the motion of vacuolar volutin granules in Saccharomyces cerevisiae cells. The motion of granules is a non-Markovian process. It does not depend on the cell cycle phase, but depends on the growth stage. The maximal number of cells with "dancing bodies" was observed under cultivation of yeast at 25-28 °C and pH 5.4-5.8. Irradiation by non-ionizing electromagnetic radiation (EMR) of extremely high frequency (61.22 GHz, 100 μW, 30 min) had no effect on granule motion. After irradiation by non-ionizing EMR of very high frequency (40.68 MHz, 30 W, 30 min) the number of cells with "dancing bodies" decreased significantly and in 2 h restored almost to the control value. The possible nature of the moving volutin granules phenomenon due to metabolic processes is discussed.
- MeSH
- elektromagnetické záření MeSH
- koncentrace vodíkových iontů MeSH
- konfokální mikroskopie MeSH
- kultivační média MeSH
- počítačové zpracování obrazu MeSH
- pohyb těles MeSH
- polyfosfáty chemie MeSH
- Saccharomyces cerevisiae chemie cytologie účinky záření MeSH
- teplota MeSH
- vakuoly chemie účinky záření MeSH
- Publikační typ
- časopisecké články MeSH
Cíl: Popsat nález neobvyklého močového konkrementu u pětiletého pacienta s kongenitální hypofosfatázií. Typ studie: Kazuistika se zhodnocením laboratorních nálezů. Název a sídlo pracoviště: Ústav lékařské chemie a klinické biochemie 2. lékařské fakulty Univerzity Karlovy a Fakultní nemocnice Motol, Pediatrická klinika 2. lékařské fakulty Univerzity Karlovy a Fakultní nemocnice Motol, V Úvalu 84, 150 06 Praha 5 Materiál a metody: Popis kazuistiky se zaměřením se především na analýzy močových konkrementů pomocí polarizační mikroskopie a infračervené spektrometrie. Výsledky: U pětiletého pacienta s kongenitální hypofosfatázií efektivně léčeného rekombinantní alkalickou fosfatázou (jejíž hodnota v séru představovala desetinásobek horní hranice referenčního rozmezí) byl prokázán raritní pyrofosfátový močový konkrement s příměsí sulfátu. K jeho tvorbě došlo v období, kdy byla u pacienta terapeuticky alkalizována moč (jejímž důvodem byla metafylaxe urátové urolitiázy), a pacient užíval profylakticky sulfonamidy před plánovanou litotrypsí. Závěr: Ačkoliv sérová koncentrace alkalické fosfatázy, jejíž funkcí je přeměna pyrofosfátu na fosfát, byla vysoká, byl u pacienta s kongenitální hypofosfatázií identifikován pyrofosfátový močový konkrement. Sulfátová složka konkrementu vznikla pravděpodobně na podkladě profylaktického podávání sulfonamidů.
Objectives: Identification of unusual renal pyrophosphate stone in 5 years old patient with congenital hypophosphatasia. Design: Case report and evaluation of laboratory results. Settings: Department of Medical Chemistry and Clinical Biochemistry, University Hospital Motol, Charles University, Second Faculty of Medicine, Department of Paediatrics, University Hospital Motol, Charles University, Second Faculty of Medicine, V Úvalu 84, 150 06 Prague 5 (Czech Republic). Material and methods: Case report focusing on determination of urinary stone composition by polarized light microscopy and infra-red spectrometry. Results: In 5 years old patient with effectively treated congenital hypophosphatasia by recombinant alkaline phosphatase (the serum value of which was ten times the upper limit of the reference range), rare pyrophosphate renal stone with addition of sulphate was demonstrated. The formation of the stone occurred in the period when the patient´s urine was therapeutically alkalized (due to the history of urate stones) and the patient used prophylactically sulfonamides prior to planned lithotripsy. Conclusion: Despite the high serum alkaline phosphatase concentration, the function of which is the conversion of pyrophosphate to phosphate, pyrophosphate renal stone was identified in a patient with congenital hypophosphatasia. The sulphate component of the stone is probably due to the prophylactic administration of sulfonamides.
- Klíčová slova
- infračervená spektrometrie,
- MeSH
- analýza moči metody MeSH
- chemické techniky analytické metody MeSH
- difosfáty analýza MeSH
- hypofosfatázie * komplikace patologie MeSH
- klinické laboratorní techniky metody MeSH
- ledvinové kameny * etiologie chemie terapie MeSH
- lidé MeSH
- litotripse metody MeSH
- předškolní dítě MeSH
- sulfonamidy terapeutické užití MeSH
- výsledek terapie MeSH
- Check Tag
- lidé MeSH
- mužské pohlaví MeSH
- předškolní dítě MeSH
- Publikační typ
- kazuistiky MeSH
- MeSH
- bezpečnost potravin MeSH
- chuťové esence MeSH
- dusitan sodný analýza normy MeSH
- geneticky modifikované potraviny MeSH
- kvalita jídla MeSH
- masné výrobky analýza normy MeSH
- označování potravin MeSH
- polyfosfáty analýza normy MeSH
- potravinářská barviva MeSH
- potravinářská technologie metody normy MeSH
- potravinářské konzervační látky analýza normy MeSH
- potravinářské přísady * MeSH
