scholarly journals The eisosome core is composed of BAR domain proteins

2011 ◽  
Vol 22 (13) ◽  
pp. 2360-2372 ◽  
Author(s):  
Agustina Olivera-Couto ◽  
Martin Graña ◽  
Laura Harispe ◽  
Pablo S. Aguilar
Keyword(s):  
Plasma Membrane ◽  
Mammalian Cells ◽  
Site Directed Mutagenesis ◽  
Bar Domain ◽  
Macromolecular Assemblies ◽  
Surface Patches ◽  
Cytoplasmic Proteins ◽  
Associated Proteins ◽  
Core Components ◽  
Membrane Bending

Eisosomes define sites of plasma membrane organization. In Saccharomyces cerevisiae, eisosomes delimit furrow-like plasma membrane invaginations that concentrate sterols, transporters, and signaling molecules. Eisosomes are static macromolecular assemblies composed of cytoplasmic proteins, most of which have no known function. In this study, we used a bioinformatics approach to analyze a set of 20 eisosome proteins. We found that the core components of eisosomes, paralogue proteins Pil1 and Lsp1, are distant homologues of membrane-sculpting Bin/amphiphysin/Rvs (BAR) proteins. Consistent with this finding, purified recombinant Pil1 and Lsp1 tubulated liposomes and formed tubules when the proteins were overexpressed in mammalian cells. Structural homology modeling and site-directed mutagenesis indicate that Pil1 positively charged surface patches are needed for membrane binding and liposome tubulation. Pil1 BAR domain mutants were defective in both eisosome assembly and plasma membrane domain organization. In addition, we found that eisosome-associated proteins Slm1 and Slm2 have F-BAR domains and that these domains are needed for targeting to furrow-like plasma membrane invaginations. Our results support a model in which BAR domain protein–mediated membrane bending leads to clustering of lipids and proteins within the plasma membrane.

Sequence Analysis of a Subset of Plasma Membrane Raft Proteome Containing CXXC Metal Binding Motifs

2015 ◽  
Vol 5 (2) ◽  
pp. 1-15
Author(s):  
Santosh Kumar Sahu ◽  
Himadri Gourav Behuria ◽  
Sangam Gupta ◽  
Babita Sahoo
Keyword(s):  
Plasma Membrane ◽  
Metal Binding ◽  
Mammalian Cells ◽  
Membrane Raft ◽  
Binding Motifs ◽  
Cxxc Motif ◽  
Signaling Mechanisms ◽  
Associated Proteins ◽  
Heavy Metal Sensing ◽  
Metal Sensing

In an attempt to identify the metal sensing proteins localized to mammalian plasma membrane, the authors screened a list of 300 raft associated proteins that are involved in cellular signaling mechanisms by searching the presence of metal thionin (CXXC) motifs. 50 proteins were found to possess CXXC motifs that could act as potential metal sensing proteins. The authors determined membrane topologies of the above CXXC motif containing proteins using TM-pred and analyzed the positions of their transmembrane (TM) domains using Bio-edit software. Based on the topology of CXXC domains, the authors classified all the raft-associated metal sensing proteins into six categories. They are (i) Exoplasmic tails with CXXC motif, (ii) Exoplasmic loops with CXXC motif, (iii) Cytosolic tails with CXXC motif, (iv) Cytosolic loop with CXXC motif, (v) TM domains with CXXC motifs, (vi) Proteins with multiple topologies of CXXC motif. The authors' study will lead to understanding of the raft-mediated mechanism of heavy metal sensing and signaling in mammalian cells.

scholarly journals An Abl-FBP17 mechanosensing system couples local plasma membrane curvature and stress fiber remodeling during mechanoadaptation

2019 ◽  
Vol 10 (1) ◽  
Author(s):  
Asier Echarri ◽  
Dácil M. Pavón ◽  
Sara Sánchez ◽  
María García-García ◽  
Enrique Calvo ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Structural Changes ◽  
Osmotic Shock ◽  
Mechanical Strain ◽  
Membrane Curvature ◽  
Bar Domain ◽  
Abl Tyrosine Kinase ◽  
Cytoskeleton Remodeling ◽  
Membrane Bending ◽  
Biochemical Signals

AbstractCells remodel their structure in response to mechanical strain. However, how mechanical forces are translated into biochemical signals that coordinate the structural changes observed at the plasma membrane (PM) and the underlying cytoskeleton during mechanoadaptation is unclear. Here, we show that PM mechanoadaptation is controlled by a tension-sensing pathway composed of c-Abl tyrosine kinase and membrane curvature regulator FBP17. FBP17 is recruited to caveolae to induce the formation of caveolar rosettes. FBP17 deficient cells have reduced rosette density, lack PM tension buffering capacity under osmotic shock, and cannot adapt to mechanical strain. Mechanistically, tension is transduced to the FBP17 F-BAR domain by direct phosphorylation mediated by c-Abl, a mechanosensitive molecule. This modification inhibits FBP17 membrane bending activity and releases FBP17-controlled inhibition of mDia1-dependent stress fibers, favoring membrane adaptation to increased tension. This mechanoprotective mechanism adapts the cell to changes in mechanical tension by coupling PM and actin cytoskeleton remodeling.

Regulation of the Actin Cytoskeleton-Plasma Membrane Interplay by Phosphoinositides

2010 ◽  
Vol 90 (1) ◽  
pp. 259-289 ◽  
Author(s):  
Juha Saarikangas ◽  
Hongxia Zhao ◽  
Pekka Lappalainen
Keyword(s):  
Plasma Membrane ◽  
Actin Cytoskeleton ◽  
Actin Dynamics ◽  
Actin Binding ◽  
Cell Physiology ◽  
Cortical Actin ◽  
Bar Domain ◽  
Pathogen Invasion ◽  
Associated Proteins ◽  
Continuous Dynamic

The plasma membrane and the underlying cortical actin cytoskeleton undergo continuous dynamic interplay that is responsible for many essential aspects of cell physiology. Polymerization of actin filaments against cellular membranes provides the force for a number of cellular processes such as migration, morphogenesis, and endocytosis. Plasma membrane phosphoinositides (especially phosphatidylinositol bis- and trisphosphates) play a central role in regulating the organization and dynamics of the actin cytoskeleton by acting as platforms for protein recruitment, by triggering signaling cascades, and by directly regulating the activities of actin-binding proteins. Furthermore, a number of actin-associated proteins, such as BAR domain proteins, are capable of directly deforming phosphoinositide-rich membranes to induce plasma membrane protrusions or invaginations. Recent studies have also provided evidence that the actin cytoskeleton-plasma membrane interactions are misregulated in a number of pathological conditions such as cancer and during pathogen invasion. Here, we summarize the wealth of knowledge on how the cortical actin cytoskeleton is regulated by phosphoinositides during various cell biological processes. We also discuss the mechanisms by which interplay between actin dynamics and certain membrane deforming proteins regulate the morphology of the plasma membrane.

scholarly journals Lipid-modified, cysteinyl-containing peptides of diverse structures are efficiently S-acylated at the plasma membrane of mammalian cells.

1996 ◽  
Vol 134 (3) ◽  
pp. 647-660 ◽  
Author(s):  
H Schroeder ◽  
R Leventis ◽  
S Shahinian ◽  
P A Walton ◽  
J R Silvius
Keyword(s):  
Plasma Membrane ◽  
Golgi Complex ◽  
Mammalian Cells ◽  
Spectrometric Analysis ◽  
Acylation Reaction ◽  
Preferential Localization ◽  
Membrane Compartment ◽  
Associated Proteins ◽  
Modified Peptides ◽  
Almost All

A variety of cysteine-containing, lipid-modified peptides are found to be S-acylated by cultured mammalian cells. The acylation reaction is highly specific for cysteinyl over serinyl residues and for lipid-modified peptides over hydrophilic peptides. The S-acylation process appears by various criteria to be enzymatic and resembles the S-acylation of plasma membrane-associated proteins in various characteristics, including inhibition by tunicamycin. The substrate range of the S-acylation reaction encompasses, but is not limited to, lipopeptides incorporating the motifs myristoylGC- and -CXC(farnesyl)-OCH3, which are reversibly S-acylated in various intracellular proteins. Mass-spectrometric analysis indicates that palmitoyl residues constitute the predominant but not the only type of S-acyl group coupled to a lipopeptide carrying the myristoylGC- motif, with smaller amounts of S-stearoyl and S-oleoyl substituents also detectable. Fluorescence microscopy using NBD-labeled cysteinyl lipopeptides reveals that the products of lipopeptide S-acylation, which cannot diffuse between membranes, are in almost all cases localized preferentially to the plasma membrane. This preferential localization is found even at reduced temperatures where vesicular transport from the Golgi complex to the plasma membrane is suppressed, strongly suggesting that the plasma membrane itself is the preferred site of S-acylation of these species. Uniquely among the lipopeptides studied, species incorporating an unphysiological N-myristoylcysteinyl- motif also show substantial formation of S-acylated products in a second, intracellular compartment identified as the Golgi complex by its labeling with a fluorescent ceramide. Our results suggest that distinct S-acyltransferases exist in the Golgi complex and plasma membrane compartments and that S-acylation of motifs such as myristoylGC- occurs specifically at the plasma membrane, affording efficient targeting of cellular proteins bearing such motifs to this membrane compartment.

The role of membrane-shaping BAR domain proteins in caveolar invagination: from mechanistic insights to pathophysiological consequences

2020 ◽  
Vol 48 (1) ◽  
pp. 137-146 ◽  
Author(s):  
Michael M. Kessels ◽  
Britta Qualmann
Keyword(s):  
Plasma Membrane ◽  
Cultured Cells ◽  
Clinical Symptoms ◽  
Membrane Damage ◽  
Model Systems ◽  
Loss Of Function ◽  
Bar Domain ◽  
Core Components ◽  
And Function

The formation of caveolae, bulb-shaped plasma membrane invaginations, requires the coordinated action of distinct lipid-interacting and -shaping proteins. The interdependence of caveolar structure and function has evoked substantial scientific interest given the association of human diseases with caveolar dysfunction. Model systems deficient of core components of caveolae, caveolins or cavins, did not allow for an explicit attribution of observed functional defects to the requirement of caveolar invagination as they lack both invaginated caveolae and caveolin proteins. Knockdown studies in cultured cells and recent knockout studies in mice identified an additional family of membrane-shaping proteins crucial for caveolar formation, syndapins (PACSINs) — BAR domain superfamily proteins characterized by crescent-shaped membrane binding interfaces recognizing and inducing distinct curved membrane topologies. Importantly, syndapin loss-of-function resulted exclusively in impairment of caveolar invagination without a reduction in caveolin or cavin at the plasma membrane, thereby allowing the specific role of the caveolar invagination to be unveiled. Muscle cells of syndapin III KO mice showed severe reductions of caveolae reminiscent of human caveolinopathies and were more vulnerable to membrane damage upon changes in membrane tensions. Consistent with the lack of syndapin III-dependent invaginated caveolae providing mechanoprotection by releasing membrane reservoirs through caveolar flattening, physical exercise of syndapin III KO mice resulted in pathological defects reminiscent of the clinical symptoms of human myopathies associated with caveolin 3 mutation suggesting that the ability of muscular caveolae to respond to mechanical forces is a key physiological process.

Biotinylation and Purification of Plasma Membrane-associated Proteins from Rodent Cultured Neurons

BIO-PROTOCOL ◽  
2016 ◽  
Vol 6 (10) ◽  
Author(s):  
Margarida Caldeira ◽  
Joana Ferreira ◽  
Ana Carvalho ◽  
Carlos Duarte
Keyword(s):  
Plasma Membrane ◽  
Cultured Neurons ◽  
Associated Proteins

scholarly journals Hakai is required for stabilization of core components of the m6A mRNA methylation machinery

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Praveen Bawankar ◽  
Tina Lence ◽  
Chiara Paolantoni ◽  
Irmgard U. Haussmann ◽  
Migle Kazlauskiene ◽  
...  
Keyword(s):  
Catalytic Activity ◽  
Sex Determination ◽  
Ubiquitin Ligase ◽  
Mammalian Cells ◽  
E3 Ubiquitin Ligase ◽  
Human Cells ◽  
Biological Functions ◽  
Mrna Metabolism ◽  
Mrna Methylation ◽  
Core Components

AbstractN6-methyladenosine (m6A) is the most abundant internal modification on mRNA which influences most steps of mRNA metabolism and is involved in several biological functions. The E3 ubiquitin ligase Hakai was previously found in complex with components of the m6A methylation machinery in plants and mammalian cells but its precise function remained to be investigated. Here we show that Hakai is a conserved component of the methyltransferase complex in Drosophila and human cells. In Drosophila, its depletion results in reduced m6A levels and altered m6A-dependent functions including sex determination. We show that its ubiquitination domain is required for dimerization and interaction with other members of the m6A machinery, while its catalytic activity is dispensable. Finally, we demonstrate that the loss of Hakai destabilizes several subunits of the methyltransferase complex, resulting in impaired m6A deposition. Our work adds functional and molecular insights into the mechanism of the m6A mRNA writer complex.

scholarly journals PBRM1 Cooperates with YTHDF2 to Control HIF-1α Protein Translation

Cells ◽  
2021 ◽  
Vol 10 (6) ◽  
pp. 1425
Author(s):  
Alena Shmakova ◽  
Mark Frost ◽  
Michael Batie ◽  
Niall S. Kenneth ◽  
Sonia Rocha
Keyword(s):  
Protein Expression ◽  
Cellular Response ◽  
Transcriptional Activity ◽  
Mrna Translation ◽  
Protein Translation ◽  
Signalling Pathways ◽  
Independent Function ◽  
Protein Levels ◽  
Associated Proteins ◽  
Core Components

PBRM1, a component of the chromatin remodeller SWI/SNF, is often deleted or mutated in human cancers, most prominently in renal cancers. Core components of the SWI/SNF complex have been shown to be important for the cellular response to hypoxia. Here, we investigated how PBRM1 controls HIF-1α activity. We found that PBRM1 is required for HIF-1α transcriptional activity and protein levels. Mechanistically, PBRM1 is important for HIF-1α mRNA translation, as absence of PBRM1 results in reduced actively translating HIF-1α mRNA. Interestingly, we found that PBRM1, but not BRG1, interacts with the m6A reader protein YTHDF2. HIF-1α mRNA is m6A-modified, bound by PBRM1 and YTHDF2. PBRM1 is necessary for YTHDF2 binding to HIF-1α mRNA and reduction of YTHDF2 results in reduced HIF-1α protein expression in cells. Our results identify a SWI/SNF-independent function for PBRM1, interacting with HIF-1α mRNA and the epitranscriptome machinery. Furthermore, our results suggest that the epitranscriptome-associated proteins play a role in the control of hypoxia signalling pathways.

scholarly journals Scanning Electron-Assisted Dielectric Microscopy Reveals Autophagosome Formation by LC3 and ATG12 in Cultured Mammalian Cells

2021 ◽  
Vol 22 (4) ◽  
pp. 1834
Author(s):  
Tomoko Okada ◽  
Toshihiko Ogura
Keyword(s):  
Mammalian Cells ◽  
Culture Medium ◽  
Related Gene ◽  
Autophagosome Formation ◽  
Intact Cells ◽  
Microtubule Associated Proteins ◽  
Associated Proteins ◽  
The Difference ◽  
Related Proteins ◽  
Scanning Electron

Autophagy is an intracellular self-devouring system that plays a central role in cellular recycling. The formation of functional autophagosomes depends on several autophagy-related proteins, including the microtubule-associated proteins 1A/1B light chain 3 (LC3) and the conserved autophagy-related gene 12 (Atg12). We have recently developed a novel scanning electron-assisted dielectric microscope (SE-ADM) for nanoscale observations of intact cells. Here, we used the SE-ADM system to observe LC3- and Atg12-containing autophagosomes in cells labelled in the culture medium with antibodies conjugated to colloidal gold particles. We observed that, during autophagosome formation, Atg12 localized along the actin meshwork structure, whereas LC3 formed arcuate or circular alignments. Our system also showed a difference in the distribution of LC3 and Atg12; Atg12 was broadly distributed while LC3 was more localized. The difference in the spatial distribution demonstrated by our system explains the difference in the size of fluorescent spots due to the fluorescently labelled antibodies observed using optical microscopy. The direct SE-ADM observation of cells should thus be effective in analyses of autophagosome formation.

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