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

Daptomycin is a calcium-dependent lipodepsipeptide antibiotic clinically used to treat serious infections caused by Gram-positive pathogens. Its precise mode of action is somewhat controversial; the biggest issue is daptomycin pore formation, which we directly investigated here. We first performed a screening experiment using propidium iodide (PI) entry to Bacillus subtilis cells and chose the optimum and therapeutically relevant conditions (10 µg/ml daptomycin and 1.25 mM CaCl2) for the subsequent analyses. Using conductance measurements on planar lipid bilayers, we show that daptomycin forms nonuniform oligomeric pores with conductance ranging from 120 pS to 14 nS. The smallest conductance unit is probably a dimer; however, tetramers and pentamers occur in the membrane most frequently. Moreover, daptomycin pore-forming activity is exponentially dependent on the applied membrane voltage. We further analyzed the membrane-permeabilizing activity in B. subtilis cells using fluorescence methods [PI and DiSC3(5)]. Daptomycin most rapidly permeabilizes cells with high initial membrane potential and dissipates it within a few minutes. Low initial membrane potential hinders daptomycin pore formation.

• Klíčová slova
• Bacillus subtilis, Staphylococcus aureus, antimicrobial lipopeptides, daptomycin, membrane, pore formation, single pore conductance,
• MeSH
• antibakteriální látky farmakologie   MeSH
• Bacillus subtilis účinky léků   metabolismus   MeSH
• biologický transport fyziologie   MeSH
• cytotoxické proteiny tvořící póry farmakologie   MeSH
• daptomycin farmakologie   MeSH
• membránové potenciály účinky léků   MeSH
• mikrobiální testy citlivosti MeSH
• permeabilita buněčné membrány účinky léků   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• antibakteriální látky MeSH
• cytotoxické proteiny tvořící póry MeSH
• daptomycin MeSH

Ever since technologies enabled the characterization of eukaryotic plasma membranes, heterogeneities in the distributions of its constituents were observed. Over the years this led to the proposal of various models describing the plasma membrane organization such as lipid shells, picket-and-fences, lipid rafts, or protein islands, as addressed in numerous publications and reviews. Instead of emphasizing on one model we in this review give a brief overview over current models and highlight how current experimental work in one or the other way do not support the existence of a single overarching model. Instead, we highlight the vast variety of membrane properties and components, their influences and impacts. We believe that highlighting such controversial discoveries will stimulate unbiased research on plasma membrane organization and functionality, leading to a better understanding of this essential cellular structure.

• Klíčová slova
• heterogenous distribution, membrane organization models, membrane physical properties, nanodomains, plasma membrane,
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH