Biological membranes play a crucial role in actively hosting, modulating and coordinating a wide range of molecular events essential for cellular function. Membranes are organized into diverse domains giving rise to dynamic molecular patchworks. However, the very definition of membrane domains has been the subject of continuous debate. For example, in the plant field, membrane domains are often referred to as nanodomains, nanoclusters, microdomains, lipid rafts, membrane rafts, signalling platforms, foci or liquid-ordered membranes without any clear rationale. In the context of plant-microbe interactions, microdomains have sometimes been used to refer to the large area at the plant-microbe interface. Some of these terms have partially overlapping meanings at best, but they are often used interchangeably in the literature. This situation generates much confusion and limits conceptual progress. There is thus an urgent need for us as a scientific community to resolve these semantic and conceptual controversies by defining an unambiguous nomenclature of membrane domains. In this Review, experts in the field get together to provide explicit definitions of plasma membrane domains in plant systems and experimental guidelines for their study. We propose that plasma membrane domains should not be considered on the basis of their size alone but rather according to the biological system being considered, such as the local membrane environment or the entire cell.

• MeSH
• buněčná membrána * metabolismus   MeSH
• membránové mikrodomény * metabolismus   MeSH
• rostliny * MeSH
• terminologie jako téma * MeSH
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH

Fluorescence light microscopy provided convincing evidence for the domain organization of plant plasma membrane (PM) proteins. Both peripheral and integral PM proteins show an inhomogeneous distribution within the PM. However, the size of PM nanodomains and protein clusters is too small to accurately determine their dimensions and nano-organization using routine confocal fluorescence microscopy and super-resolution methods. To overcome this limitation, we have developed a novel correlative light electron microscopy method (CLEM) using total internal reflection fluorescence microscopy (TIRFM) and advanced environmental scanning electron microscopy (A-ESEM). Using this technique, we determined the number of auxin efflux carriers from the PINFORMED (PIN) family (NtPIN3b-GFP) within PM nanodomains of tobacco cell PM ghosts. Protoplasts were attached to coverslips and immunostained with anti-GFP primary antibody and secondary antibody conjugated to fluorochrome and gold nanoparticles. After imaging the nanodomains within the PM with TIRFM, the samples were imaged with A-ESEM without further processing, and quantification of the average number of molecules within the nanodomain was performed. Without requiring any post-fixation and coating procedures, this method allows to study details of the organization of auxin carriers and other plant PM proteins.

• Klíčová slova
• auxin carriers, correlative microscopy, nanodomains, plasma membrane,
• MeSH
• Arabidopsis genetika   růst a vývoj   MeSH
• buněčná membrána genetika   metabolismus   ultrastruktura   MeSH
• konfokální mikroskopie MeSH
• kovové nanočástice chemie   MeSH
• kyseliny indoloctové metabolismus   MeSH
• mikroskopie elektronová rastrovací * MeSH
• počítačové zpracování obrazu MeSH
• protoplasty metabolismus   ultrastruktura   MeSH
• regulátory růstu rostlin genetika   metabolismus   MeSH
• tabák genetika   metabolismus   ultrastruktura   MeSH
• zlato chemie   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• kyseliny indoloctové MeSH
• regulátory růstu rostlin MeSH
• zlato MeSH

Membrane proteins are targeted not only to specific membranes in the cell architecture, but also to distinct lateral microdomains within individual membranes to properly execute their biological functions. Yeast tetraspan protein Nce102 has been shown to migrate between such microdomains within the plasma membrane in response to an acute drop in sphingolipid levels. Combining microscopy and biochemistry methods, we show that upon gradual ageing of a yeast culture, when sphingolipid demand increases, Nce102 migrates from the plasma membrane to the vacuole. Instead of being targeted for degradation it localizes to V-ATPase-poor, i.e., ergosterol-enriched, domains of the vacuolar membrane, analogous to its plasma membrane localization. We discovered that, together with its homologue Fhn1, Nce102 modulates vacuolar morphology, dynamics, and physiology. Specifically, the fusing of vacuoles, accompanying a switch of fermenting yeast culture to respiration, is retarded in the strain missing both proteins. Furthermore, the absence of either causes an enlargement of ergosterol-rich vacuolar membrane domains, while the vacuoles themselves become smaller. Our results clearly show decreased stability of the V-ATPase in the absence of either Nce102 or Fhn1, a possible result of the disruption of normal microdomain morphology of the vacuolar membrane. Therefore, the functionality of the vacuole as a whole might be compromised in these cells.

• Klíčová slova
• eisosome, membrane microdomains, sphingolipid metabolism, vacuolar morphology, yeast,
• MeSH
• Saccharomyces cerevisiae - proteiny metabolismus   MeSH
• Saccharomyces cerevisiae metabolismus   MeSH
• vakuoly metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• NCE102 protein, S cerevisiae MeSH Prohlížeč
• Saccharomyces cerevisiae - proteiny MeSH

One of the best characterized fungal membrane microdomains is the MCC/eisosome. The MCC (membrane compartment of Can1) is an evolutionarily conserved ergosterol-rich plasma membrane domain. It is stabilized on its cytosolic face by the eisosome, a hemitubular protein complex composed of Bin/Amphiphysin/Rvs (BAR) domain-containing Pil1 and Lsp1. These two proteins bind directly to phosphatidylinositol 4,5-bisphosphate and promote the typical furrow-like shape of the microdomain, with highly curved edges and bottom. While some proteins display stable localization in the MCC/eisosome, others enter or leave it under particular conditions, such as misbalance in membrane lipid composition, changes in membrane tension, or availability of specific nutrients. These findings reveal that the MCC/eisosome, a plasma membrane microdomain with distinct morphology and lipid composition, acts as a multifaceted regulator of various cellular processes including metabolic pathways, cellular morphogenesis, signalling cascades, and mRNA decay. In this minireview, we focus on the MCC/eisosome's proposed role in the regulation of lipid metabolism. While the molecular mechanisms of the MCC/eisosome function are not completely understood, the idea of intracellular processes being regulated at the plasma membrane, the foremost barrier exposed to environmental challenges, is truly exciting.

• Klíčová slova
• MCC, eisosome, ergosterol, lipids, microdomain, phosphoinositides, regulation, sphingolipids,
• MeSH
• buněčná membrána metabolismus   MeSH
• fosfatidylinositol-4,5-difosfát metabolismus   MeSH
• fungální proteiny chemie   metabolismus   MeSH
• homeostáza MeSH
• houby metabolismus   MeSH
• metabolismus lipidů MeSH
• proteinové domény MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH
• Názvy látek
• fosfatidylinositol-4,5-difosfát MeSH
• fungální proteiny MeSH

Quantitative approaches for characterizing molecular organization of cell membrane molecules under physiological and pathological conditions profit from recently developed super-resolution imaging techniques. Current tools employ statistical algorithms to determine clusters of molecules based on single-molecule localization microscopy (SMLM) data. These approaches are limited by the ability of SMLM techniques to identify and localize molecules in densely populated areas and experimental conditions of sample preparation and image acquisition. We have developed a robust, model-free, quantitative clustering analysis to determine the distribution of membrane molecules that excels in densely labeled areas and is tolerant to various experimental conditions, i.e. multiple-blinking or high blinking rates. The method is based on a TIRF microscope followed by a super-resolution optical fluctuation imaging (SOFI) analysis. The effectiveness and robustness of the method is validated using simulated and experimental data investigating nanoscale distribution of CD4 glycoprotein mutants in the plasma membrane of T cells.

• MeSH
• algoritmy MeSH
• antigeny CD4 genetika   metabolismus   MeSH
• buněčná membrána metabolismus   MeSH
• fluorescenční barviva MeSH
• fluorescenční mikroskopie metody   statistika a číselné údaje   MeSH
• Jurkat buňky MeSH
• lidé MeSH
• membránové proteiny genetika   metabolismus   MeSH
• mutantní proteiny genetika   metabolismus   MeSH
• optické zobrazování metody   statistika a číselné údaje   MeSH
• shluková analýza MeSH
• T-lymfocyty imunologie   metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• validační studie MeSH
• Názvy látek
• antigeny CD4 MeSH
• fluorescenční barviva MeSH
• membránové proteiny MeSH
• mutantní proteiny MeSH

The plasma membrane of the fungal pathogen Candida albicans forms a protective barrier that also mediates many processes needed for virulence, including cell wall synthesis, invasive hyphal morphogenesis, and nutrient uptake. Because compartmentalization of the plasma membrane is believed to coordinate these diverse activities, we examined plasma membrane microdomains termed eisosomes or membrane compartment of Can1 (MCC), which correspond to ∼200-nm-long furrows in the plasma membrane. A pil1∆ lsp1∆ mutant failed to form eisosomes and displayed strong defects in plasma membrane organization and morphogenesis, including extensive cell wall invaginations. Mutation of eisosome proteins Slm2, Pkh2, and Pkh3 did not cause similar cell wall defects, although pkh2∆ cells formed chains of furrows and pkh3∆ cells formed wider furrows, identifying novel roles for the Pkh protein kinases in regulating furrows. In contrast, the sur7∆ mutant formed cell wall invaginations similar to those for the pil1∆ lsp1∆ mutant even though it could form eisosomes and furrows. A PH-domain probe revealed that the regulatory lipid phosphatidylinositol 4,5-bisphosphate was enriched at sites of cell wall invaginations in both the sur7∆ and pil1∆ lsp1∆ cells, indicating that this contributes to the defects. The sur7∆ and pil1∆ lsp1∆ mutants displayed differential susceptibility to various types of stress, indicating that they affect overlapping but distinct functions. In support of this, many mutant phenotypes of the pil1∆ lsp1∆ cells were rescued by overexpressing SUR7 These results demonstrate that C. albicans eisosomes promote the ability of Sur7 to regulate plasma membrane organization.

• MeSH
• buněčná membrána metabolismus   MeSH
• buněčná stěna metabolismus   MeSH
• Candida albicans metabolismus   MeSH
• endocytóza fyziologie   MeSH
• fosfoproteiny metabolismus   MeSH
• fungální proteiny metabolismus   MeSH
• hyfy metabolismus   MeSH
• membránové mikrodomény metabolismus   MeSH
• membránové proteiny metabolismus   MeSH
• proteinkinasy metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• fosfoproteiny MeSH
• fungální proteiny MeSH
• membránové proteiny MeSH
• proteinkinasy MeSH

Regulation of gene expression on the level of translation and mRNA turnover is widely conserved evolutionarily. We have found that the main mRNA decay enzyme, exoribonuclease Xrn1, accumulates at the plasma membrane-associated eisosomes after glucose exhaustion in a culture of the yeast S. cerevisiae. Eisosomal localization of Xrn1 is not achieved in cells lacking the main component of eisosomes, Pil1, or Sur7, the protein accumulating at the membrane compartment of Can1 (MCC) - the eisosome-organized plasma membrane microdomain. In contrast to the conditions of diauxic shift, when Xrn1 accumulates in processing bodies (P-bodies), or acute heat stress, in which these cytosolic accumulations of Xrn1 associate with eIF3a/Rpg1-containing stress granules, Xrn1 is not accompanied by other mRNA-decay machinery components when it accumulates at eisosomes in post-diauxic cells. It is important that Xrn1 is released from eisosomes after addition of fermentable substrate. We suggest that this spatial segregation of Xrn1 from the rest of the mRNA-decay machinery reflects a general regulatory mechanism, in which the key enzyme is kept separate from the rest of mRNA decay factors in resting cells but ready for immediate use when fermentable nutrients emerge and appropriate metabolism reprogramming is required. In particular, the localization of Xrn1 to the eisosome, together with previously published data, accents the relevance of this plasma membrane-associated compartment as a multipotent regulatory site.

• MeSH
• buněčná membrána genetika   metabolismus   MeSH
• exoribonukleasy genetika   metabolismus   MeSH
• exprese genu MeSH
• glukosa metabolismus   MeSH
• reakce na tepelný šok MeSH
• rekombinantní fúzní proteiny genetika   metabolismus   MeSH
• reportérové geny MeSH
• Saccharomyces cerevisiae - proteiny genetika   metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• exoribonukleasy MeSH
• glukosa MeSH
• rekombinantní fúzní proteiny MeSH
• Saccharomyces cerevisiae - proteiny MeSH
• XRN1 protein, S cerevisiae MeSH Prohlížeč