Article ID | Journal | Published Year | Pages | File Type |
---|---|---|---|---|
8469613 | European Journal of Cell Biology | 2018 | 10 Pages |
Abstract
In budding yeast Saccharomyces cerevisiae, PtdIns(4)P serves as an essential signalling molecule in the Golgi complex, endosomal system, and plasma membrane, where it is involved in the control of multiple cellular functions via direct interactions with PtdIns(4)P-binding proteins. To analyse the distribution of PtdIns(4)P in yeast cells at a nanoscale level, we employed an electron microscopy technique that specifically labels PtdIns(4)P on the freeze-fracture replica of the yeast membrane. This method minimizes the possibility of artificial perturbation, because molecules in the membrane are physically immobilised in situ. We observed that PtdIns(4)P is localised on the cytoplasmic leaflet, but not the exoplasmic leaflet, of the plasma membrane, Golgi body, vacuole, and vesicular structure membranes. PtdIns(4)P labelling was not observed in the membrane of the endoplasmic reticulum, and in the outer and inner membranes of the nuclear envelope or mitochondria. PtdIns(4)P forms clusters of <100â¯nm in diameter in the plasma membrane and vacuolar membrane according to point pattern analysis of immunogold labelling. There are three kinds of compartments in the cytoplasmic leaflet of the plasma membrane. In the present study, we showed that PtdIns(4)P is specifically localised in the flat undifferentiated plasma membrane compartment. In the vacuolar membrane, PtdIns(4)P was concentrated in intramembrane particle (IMP)-deficient raft-like domains, which are tightly bound to lipid droplets, but not surrounding IMP-rich non-raft domains in geometrical IMP-distributed patterns in the stationary phase. This is the first report showing microdomain formations of PtdIns(4)P in the plasma membrane and vacuolar membrane of budding yeast cells at a nanoscale level, which will illuminate the functionality of PtdIns(4)P in each membrane.
Keywords
PLCPPINintramembrane particleOCRLPtdInsOSBPIMPMCCMCTFRAPGSTSDSMCPCSRGFPComplete spatial randomnessLTPVacuolesodium dodecylsulfateFreeze-fracturePlasma membranephosphatase and tensin homolog deleted from chromosome 10phosphatidylinositolPhosphoinositidesphospholipase Cfluorescence recovery after photobleachinglipid dropletLipidYeastElectron microscopyPleckstrin HomologyOxysterol-binding proteinlipid transfer proteingreen fluorescent proteinPtenglutathione S-transferase
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Authors
Kan-na Tomioku, Mikiko Shigekuni, Hiroki Hayashi, Akane Yoshida, Taiki Futagami, Hisanori Tamaki, Kenji Tanabe, Akikazu Fujita,