Information about cholesterol subcellular localization and transport pathways inside cells is essential for understanding and treatment of cholesterol-related diseases. However, there is a lack of reliable tools to monitor it. This work follows the fate of Sterolight, a BODIPY-labelled sterol, within the cell and demonstrates it as a suitable probe for visualization of sterol/lipid trafficking. Sterolight enters cells through an energy-independent process and knockdown experiments suggest caveolin-1 as its potential cellular carrier. Intracellular transport of Sterolight is a rapid process, and transfer from ER and mitochondria to lysosomes and later to lipid droplets requires the participation of active microtubules, as it can be inhibited by the microtubule disruptor nocodazole. Excess of the probe is actively exported from cells, in addition to being stored in lipid droplets, to re-establish the sterol balance. Efflux occurs through a mechanism requiring energy and may be selectively poisoned with verapamil or blocked in cells with mutated cholesterol transporter NPC1. Sterolight is efficiently transferred within and between different cell populations, making it suitable for monitoring numerous aspects of sterol biology, including the live tracking and visualization of intracellular and intercellular transport.
- MeSH
- cholesterol krev MeSH
- desmosterol krev MeSH
- hypercholesterolemie * diagnóza MeSH
- lidé MeSH
- pilotní projekty MeSH
- retrospektivní studie MeSH
- sitosteroly krev MeSH
- steroly klasifikace metabolismus MeSH
- těhotenství * metabolismus MeSH
- Check Tag
- lidé MeSH
- těhotenství * metabolismus MeSH
- ženské pohlaví MeSH
- MeSH
- cholesterol izolace a purifikace metabolismus MeSH
- desmosterol izolace a purifikace metabolismus MeSH
- HDL-cholesterol izolace a purifikace metabolismus MeSH
- hypercholesterolemie * diagnóza etnologie MeSH
- lanosterol izolace a purifikace metabolismus MeSH
- LDL-cholesterol izolace a purifikace metabolismus MeSH
- lidé MeSH
- retrospektivní studie * MeSH
- sitosteroly izolace a purifikace metabolismus MeSH
- statistika jako téma MeSH
- steroly izolace a purifikace metabolismus MeSH
- těhotenství * fyziologie metabolismus MeSH
- triglyceridy izolace a purifikace metabolismus MeSH
- Check Tag
- lidé MeSH
- těhotenství * fyziologie metabolismus MeSH
- ženské pohlaví MeSH
- MeSH
- Alzheimerova nemoc * metabolismus patologie MeSH
- antiflogistika nesteroidní terapeutické užití MeSH
- apolipoproteiny metabolismus MeSH
- buněčná membrána metabolismus MeSH
- centrální nervový systém metabolismus MeSH
- cholesterol metabolismus MeSH
- fosfolipidy metabolismus MeSH
- glycerofosfolipidy metabolismus MeSH
- kanabinoidy metabolismus MeSH
- kyselina arachidonová MeSH
- kyseliny dokosahexaenové terapeutické užití MeSH
- lidé MeSH
- lipoproteiny metabolismus MeSH
- mastné kyseliny terapeutické užití MeSH
- mozek metabolismus účinky léků MeSH
- scavengerové receptory - třída B metabolismus MeSH
- sfingolipidy metabolismus MeSH
- steroly metabolismus MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- práce podpořená grantem MeSH
- Klíčová slova
- oxidační produkty sterolů,
- MeSH
- cholesterol * chemie metabolismus škodlivé účinky MeSH
- fytosteroly * chemie metabolismus škodlivé účinky MeSH
- ketocholesteroly * analýza chemie metabolismus škodlivé účinky MeSH
- lidé MeSH
- maso * analýza škodlivé účinky využití MeSH
- oxidace-redukce * MeSH
- oxysteroly MeSH
- potraviny * MeSH
- statistika jako téma * MeSH
- steroly * chemie metabolismus škodlivé účinky MeSH
- technologie * metody normy organizace a řízení trendy MeSH
- vysoká teplota * škodlivé účinky MeSH
- Check Tag
- lidé MeSH
Ectotherm animals including insects are known to undergo seasonal restructuring of the cell membranes in order to keep their functionality and/or protect their structural integrity at low body temperatures. Studies on insects so far focused either on fatty acids or on composition of molecular species in major phospholipid classes. Here we extend the scope of analysis and bring results on seasonal changes in minor phospholipid classes, lysophospholipids (LPLs), free fatty acids, phytosterols and tocopherols in heteropteran insect, Pyrrhocoris apterus. We found that muscle tissue contains unusually high amounts of LPLs. Muscle and fat body tissues also contain high amounts of β-sitosterol and campesterol, two phytosterols derived from plant food, while only small amounts of cholesterol are present. In addition, two isomers (γ and δ) of tocopherol (vitamin E) are present in quantities comparable to, or even higher than phytosterols in both tissues. Distinct seasonal patterns of sterol and tocopherol concentrations were observed showing a minimum in reproductively active bugs in summer and a maximum in diapausing, cold-acclimated bugs in winter. Possible adaptive meanings of such changes are discussed including: preventing the unregulated transition of membrane lipids from functional liquid crystalline phase to non-functional gel phase; decreasing the rates of ion/solute leakage; silencing the activities of membrane bound enzymes and receptors; and counteracting the higher risk of oxidative damage to PUFA in winter membranes.
- MeSH
- buněčná membrána metabolismus MeSH
- fosfolipidy metabolismus MeSH
- Heteroptera metabolismus MeSH
- kyseliny mastné neesterifikované metabolismus MeSH
- roční období * MeSH
- steroly metabolismus MeSH
- tokoferoly metabolismus MeSH
- zvířata MeSH
- Check Tag
- mužské pohlaví MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Cryptogein is a proteinaceous elicitor secreted by Phytophthora cryptogea that can induce resistance to P. parasitica in tobacco plants. On the basis of previous computer modelling experiments, by site-directed mutagenesis a series of cryptogein variants was prepared with altered abilities to bind sterols, phospholipids or both. The sterol binding and phospholipid transfer activities corresponded well with the previously reported structural data. Induction of the synthesis of reactive oxygen species (ROS) in tobacco cells in suspension and proteomic analysis of intercellular fluid changes in tobacco leaves triggered by these mutant proteins were not proportional to their ability to bind or transfer sterols and phospholipids. However, changes in the intercellular proteome corresponded to transcription levels of defence genes and resistance to P. parasitica and structure-prediction of mutants did not reveal any significant changes in protein structure. These results suggest, contrary to previous proposals, that the sterol-binding ability of cryptogein and its mutants, and the associated conformational change in the ω-loop, might not be principal factors in either ROS production or resistance induction. Nevertheless, the results support the importance of the ω-loop for the interaction of the protein with the high affinity binding site on the plasma membrane.
- MeSH
- fosfolipidy metabolismus MeSH
- fungální proteiny genetika metabolismus MeSH
- imunita rostlin imunologie MeSH
- kyselina chlorogenová analýza MeSH
- listy rostlin genetika imunologie parazitologie fyziologie MeSH
- mutace MeSH
- nemoci rostlin imunologie parazitologie MeSH
- Phytophthora genetika metabolismus patogenita MeSH
- proteiny genetika metabolismus MeSH
- proteomika metody MeSH
- reaktivní formy kyslíku metabolismus MeSH
- rekombinantní proteiny MeSH
- seskviterpeny analýza MeSH
- steroly metabolismus MeSH
- tabák genetika imunologie parazitologie fyziologie MeSH
- vazba proteinů MeSH
- vztahy mezi strukturou a aktivitou MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- MeSH
- antifungální látky * MeSH
- buněčná membrána * metabolismus MeSH
- ergosterol MeSH
- fosfatidylethanolaminy MeSH
- fosfolipidy metabolismus MeSH
- imunitní systém * MeSH
- kardiolipiny metabolismus MeSH
- kvasinky * klasifikace MeSH
- lidé MeSH
- metabolismus lipidů * MeSH
- Saccharomyces cerevisiae MeSH
- sfingolipidy metabolismus MeSH
- steroly metabolismus MeSH
- xenobiotika * MeSH
- Check Tag
- lidé MeSH
- MeSH
- cholesterol izolace a purifikace metabolismus MeSH
- cytokiny izolace a purifikace metabolismus MeSH
- financování organizované MeSH
- granulocyty cytologie enzymologie MeSH
- hydrokortison biosyntéza metabolismus nedostatek MeSH
- hypolipoproteinemie diagnóza enzymologie metabolismus MeSH
- lidé MeSH
- nadledviny enzymologie MeSH
- polytrauma diagnóza enzymologie klasifikace MeSH
- prospektivní studie MeSH
- statistika jako téma MeSH
- steroly biosyntéza metabolismus MeSH
- syndrom systémové zánětlivé reakce diagnóza enzymologie MeSH
- Check Tag
- lidé MeSH
