scholarly journals Extensive sphingolipid depletion does not affect lipid raft integrity or lipid raft localization and efflux function of the ABC transporter MRP1

2010 ◽  
Vol 430 (3) ◽  
pp. 519-529 ◽  
Author(s):  
Karin Klappe ◽  
Anne-Jan Dijkhuis ◽  
Ina Hummel ◽  
Annie van Dam ◽  
Pavlina T. Ivanova ◽  
...  
Keyword(s):  
Fatty Acid ◽  
Lipid Rafts ◽  
Cell Lines ◽  
Lipid Raft ◽  
Related Protein ◽  
Gradient Distribution ◽  
Caveolin 1 ◽  
Free Lipid ◽  
Detergent Resistant Membranes ◽  
Multidrug Resistance Related Protein

We show that highly efficient depletion of sphingolipids in two different cell lines does not abrogate the ability to isolate Lubrol-based DRMs (detergent-resistant membranes) or detergent-free lipid rafts from these cells. Compared with control, DRM/detergent-free lipid raft fractions contain equal amounts of protein, cholesterol and phospholipid, whereas the classical DRM/lipid raft markers Src, caveolin-1 and flotillin display the same gradient distribution. DRMs/detergent-free lipid rafts themselves are severely depleted of sphingolipids. The fatty acid profile of the remaining sphingolipids as well as that of the glycerophospholipids shows several differences compared with control, most prominently an increase in highly saturated C16 species. The glycerophospholipid headgroup composition is unchanged in sphingolipid-depleted cells and cell-derived detergent-free lipid rafts. Sphingolipid depletion does not alter the localization of MRP1 (multidrug-resistance-related protein 1) in DRMs/detergent-free lipid rafts or MRP1-mediated efflux of carboxyfluorescein. We conclude that extensive sphingolipid depletion does not affect lipid raft integrity in two cell lines and does not affect the function of the lipid-raft-associated protein MRP1.

Free fatty acid transport across adipocytes is mediated by an unknown membrane protein pump

2007 ◽  
Vol 293 (5) ◽  
pp. E1207-E1214 ◽  
Author(s):  
J. Patrick Kampf ◽  
Danielle Parmley ◽  
Alan M. Kleinfeld
Keyword(s):  
Fatty Acid ◽  
Membrane Protein ◽  
Lipid Rafts ◽  
Cell Lines ◽  
Fatty Acid Transport ◽  
Acid Transport ◽  
Long Chain Fatty Acid ◽  
Caveolin 1 ◽  
Long Chain

The role of cell membranes in regulating the flux of long chain free fatty acids (FFA) into and out of adipocytes is intensely debated. Four different membrane proteins including, FABPpm, CD36/FAT, caveolin-1, and FATP have been identified as facilitating FFA transport. Moreover, CD36 and caveolin-1 are also reported to mediate transport in conjunction with lipid rafts. The principal evidence for these findings is a correlation of the level of FFA uptake with the expression level of these proteins and with the integrity of lipid rafts. The 3T3-L1 and 3T3-F442A cell lines in their preadipocyte states reveal little or no expression of these proteins and correspondingly low levels of uptake. Here we have microinjected the adipocyte and preadipocyte cell lines with ADIFAB, the fluorescent indicator of FFA. The ADIFAB fluorescence allowed us to monitor the intracellular unbound FFA concentration during FFA influx and efflux. We show that these measurements of transport, in contrast to FFA uptake measurements, correlate neither with expression of these proteins nor with lipid raft integrity in preadipocytes and adipocytes. Transport characteristics, including the generation of an ATP-dependent FFA concentration gradient, are virtually identical in adipocytes and preadipocytes. We suggest that the origin of the discrepancy between uptake and our measurements is that most of the FFA transported into the cells is lost during the uptake but not in the transport protocols. We conclude that long chain fatty acid transport in adipocytes is very likely mediated by an as-yet-unidentified membrane protein pump.

scholarly journals Lipid rafts mediate internalization of β1-integrin in migrating intestinal epithelial cells

2008 ◽  
Vol 295 (5) ◽  
pp. G965-G976 ◽  
Author(s):  
Elena V. Vassilieva ◽  
Kirsten Gerner-Smidt ◽  
Andrei I. Ivanov ◽  
Asma Nusrat
Keyword(s):  
Epithelial Cells ◽  
Lipid Rafts ◽  
Lipid Raft ◽  
Intestinal Epithelial Cells ◽  
Focal Adhesions ◽  
Endocytic Pathway ◽  
Dependent Manner ◽  
Intestinal Epithelial ◽  
Caveolin 1 ◽  
Cell Matrix

Intestinal mucosal inflammation is associated with epithelial wounds that rapidly reseal by migration of intestinal epithelial cells (IECs). Cell migration involves cycles of cell-matrix adhesion/deadhesion that is mediated by dynamic turnover (assembly and disassembly) of integrin-based focal adhesions. Integrin endocytosis appears to be critical for deadhesion of motile cells. However, mechanisms of integrin internalization during remodeling of focal adhesions of migrating IECs are not understood. This study was designed to define the endocytic pathway that mediates internalization of β1-integrin in migrating model IECs. We observed that, in SK-CO15 and T84 colonic epithelial cells, β1-integrin is internalized in a dynamin-dependent manner. Pharmacological inhibition of clathrin-mediated endocytosis or macropinocytosis and small-interfering RNA (siRNA)-mediated knock down of clathrin did not prevent β1-integrin internalization. However, β1-integrin internalization was inhibited following cholesterol extraction and after overexpression of lipid raft protein, caveolin-1. Furthermore, internalized β1-integrin colocalized with the lipid rafts marker cholera toxin, and siRNA-mediated knockdown of caveolin-1 and flotillin-1/2 increased β1-integrin endocytosis. Our data suggest that, in migrating IEC, β1-integrin is internalized via a dynamin-dependent lipid raft-mediated pathway. Such endocytosis is likely to be important for disassembly of integrin-based cell-matrix adhesions and therefore in regulating IEC migration and wound closure.

scholarly journals Caveolin-1 interacts with a lipid raft-associated population of fatty acid synthase

Cell Cycle ◽  
2008 ◽  
Vol 7 (14) ◽  
pp. 2257-2267 ◽  
Author(s):  
Dolores Di Vizio ◽  
Rosalyn M. Adam ◽  
Jayoung Kim ◽  
Robert Kim ◽  
Federica Sotgia ◽  
...  
Keyword(s):  
Fatty Acid ◽  
Lipid Raft ◽  
Fatty Acid Synthase ◽  
Caveolin 1

scholarly journals Sesquiterpene Alcohol Cedrol Chemosensitizes Human Cancer Cells and Suppresses Cell Proliferation by Destabilizing Plasma Membrane Lipid Rafts

2021 ◽  
Vol 8 ◽  
Author(s):  
Siddhartha Kumar Mishra ◽  
Yun Soo Bae ◽  
Yong-Moon Lee ◽  
Jae-Sung Kim ◽  
Seung Hyun Oh ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Cancer Cells ◽  
Lipid Rafts ◽  
Cell Lines ◽  
Lipid Raft ◽  
Human Cancer ◽  
Membrane Lipid ◽  
Human Leukemia ◽  
Sesquiterpene Alcohol ◽  
Apoptosis Protein

Chemosensitization of cancer cells with small molecules may improve the therapeutic index of antitumoral agents by making tumor cells sensitive to the drug regimen and thus overcome the treatment resistance and side effects of single therapy. Cell membrane lipid rafts are known to transduce various signaling events in cell proliferation. Sensitizing cancer cells may cause modulation of membrane lipid rafts which may potentially be used in improving anticancer drug response. Cedrol, a natural sesquiterpene alcohol, was used to treat human leukemia K562 and colon cancer HT-29 cell lines, and effects were observed. Cedrol decreased the cell viability by inducing apoptosis in both cell lines by activation of pro-apoptosis protein BID and inhibition of anti-apoptosis proteins Bcl-XL, Bcl-2, and XIAP. Cedrol activated the caspase-9-dependent mitochondrial intrinsic pathway of apoptosis. Furthermore, cedrol inhibited the levels of pAKT, pERK, and pmTOR proteins as well as nuclear and cytoplasmic levels of the p65 subunit of NF-κB. Cedrol caused redistribution of cholesterol and sphingomyelin contents from membrane lipid raft, which was confirmed by a combined additive effect with methyl-β-cyclodextrin (lipid raft-disrupting agent). Lipid raft destabilization by cedrol led to the increased production of ceramides and inhibition of membrane-bound NADPH oxidase 2 enzyme activity. Cholesterol/sphingomyelin-redistributing abilities of cedrol appear as a novel mechanism of growth inhibition of cancer cells. Cedrol can be classified as a natural lipid raft-disrupting agent with possibilities to be used in general studies involving membrane lipid raft modifications.

scholarly journals The impact of plasma membrane lipid composition on flagella-mediated adhesion of enterohemorrhagic Escherichia coli

2020 ◽  
Author(s):  
Hélène Cazzola ◽  
Laurine Lemaire ◽  
Sébastien Acket ◽  
Elise Prost ◽  
Luminita Duma ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Plasma Membrane ◽  
Fatty Acid ◽  
Linolenic Acid ◽  
Lipid Rafts ◽  
Cell Lines ◽  
Polyunsaturated Fatty Acid ◽  
Bacterial Adhesion ◽  
Human Pathogen ◽  
The Impact

AbstractEnterohemorrhagic Escherichia coli (EHEC) O157:H7 is a major cause of foodborne gastrointestinal illness. The adhesion of EHEC on host tissues is the first step enabling bacterial colonization. Adhesins like fimbriae and flagella mediate this mechanism. Here, we studied the interaction of the bacterial flagellum with the host cell’s plasma membrane using Giant Unilamellar Vesicles (GUVs) as a biologically relevant model. Cultured cell lines contain many different molecular components including proteins and glycoproteins. In contrast, with GUVs we can characterize the bacterial mode of interaction solely with a defined lipid part of the cell membrane. Bacterial adhesion on GUVs was dependent on the presence of the flagellar filament and its motility. By testing different phospholipid head groups, the nature of the fatty acid chains or the liposome curvature, we found that lipid packing is a key parameter to enable bacterial adhesion. Using HT-29 cells grown in the presence of polyunsaturated fatty acid (α-linolenic acid) or saturated fatty acid (palmitic acid), we found that α-linolenic acid reduced adhesion of wild type EHEC but not of a non-flagellated mutant. Finally, our results reveal that the presence of flagella is advantageous for the bacteria to bind to lipid rafts. We speculate that polyunsaturated fatty acids prevent flagellar adhesion on membrane bilayers and play a clear role for optimal host colonization. Flagella-mediated adhesion to plasma membranes has broad implications to host-pathogen interactions.ImportanceBacterial adhesion is a crucial step to allow bacteria to colonize their hosts, invade tissues and form biofilm. Enterohemorrhagic E. coli O157:H7 is a human pathogen and the causative agent of diarrhea and hemorrhagic colitis. Here, we use biomimetic membrane models and cell lines to decipher the impact of lipid content of the plasma membrane on enterohemorrhagic E. coli flagella-mediated adhesion. Our findings provide evidence that polyunsaturated fatty acid (α-linolenic acid) inhibits E. coli flagella adhesion to the plasma membrane in a mechanism separate from its antimicrobial and anti-inflammatory functions. In addition, we confirm that cholesterol-enriched lipid microdomains, often called lipid rafts are important in bacterial adhesion. These findings significantly strengthen plasma membrane adhesion via bacterial flagella in an important human pathogen. This mechanism represents a promising target for the development of novel anti-adhesion therapies.

CAVEOLIN-1 INTERACTS WITH A LIPID RAFT-ASSOCIATED POPULATION OF FATTY ACID SYNTHASE IN PROSTATE CANCER

2008 ◽  
Vol 179 (4S) ◽  
pp. 456-457
Author(s):  
Dolores Di Vizio ◽  
Rosalyn M Adam ◽  
Jayoung Kim ◽  
Keith R Solomon ◽  
Robert Kim ◽  
...  
Keyword(s):  
Prostate Cancer ◽  
Fatty Acid ◽  
Lipid Raft ◽  
Fatty Acid Synthase ◽  
Caveolin 1

scholarly journals Cholesterol-sensitive Modulation of Transcytosis

2007 ◽  
Vol 18 (6) ◽  
pp. 2057-2071 ◽  
Author(s):  
Julieta Leyt ◽  
Naomi Melamed-Book ◽  
Jean-Pierre Vaerman ◽  
Shulamit Cohen ◽  
Aryeh M. Weiss ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Lipid Rafts ◽  
Receptor Internalization ◽  
Cholesterol Depletion ◽  
Compensatory Mechanism ◽  
Membrane Domains ◽  
Caveolin 1 ◽  
Human Transferrin Receptor ◽  
Detergent Resistant Membranes ◽  
Tyrosine 14

Cholesterol-rich membrane domains (e.g., lipid rafts) are thought to act as molecular sorting machines, capable of coordinating the organization of signal transduction pathways within limited regions of the plasma membrane and organelles. The significance of these domains in polarized postendocytic sorting is currently not understood. We show that dimeric IgA stimulates the incorporation of its receptor into cholesterol-sensitive detergent-resistant membranes confined to the basolateral surface/basolateral endosomes. A fraction of human transferrin receptor was also found in basolateral detergent-resistant membranes. Disrupting these membrane domains by cholesterol depletion (using methyl-β-cyclodextrin) before ligand-receptor internalization caused depolarization of traffic from endosomes, suggesting that cholesterol in basolateral lipid rafts plays a role in polarized sorting after endocytosis. In contrast, cholesterol depletion performed after ligand internalization stimulated cargo transcytosis. It also stimulated caveolin-1 phosphorylation on tyrosine 14 and the appearance of the activated protein in dimeric IgA-containing apical organelles. We propose that cholesterol depletion stimulates the coupling of transcytotic and caveolin-1 signaling pathways, consequently prompting the membranes to shuttle from endosomes to the plasma membrane. This process may represent a unique compensatory mechanism required to maintain cholesterol balance on the cell surface of polarized epithelia.

scholarly journals The Sprouty-related protein, Spred-1, localizes in a lipid raft/caveola and inhibits ERK activation in collaboration with caveolin-1

2005 ◽  
Vol 10 (9) ◽  
pp. 887-895 ◽  
Author(s):  
Atsushi Nonami ◽  
Takaharu Taketomi ◽  
Akiko Kimura ◽  
Kazuko Saeki ◽  
Hiromi Takaki ◽  
...  
Keyword(s):  
Lipid Raft ◽  
Related Protein ◽  
Erk Activation ◽  
Caveolin 1

scholarly journals Synergism of rMV-Hu191 with cisplatin to treat gastric cancer by acid sphingomyelinase-mediated apoptosis requiring integrity of lipid raft microdomains

2021 ◽  
Author(s):  
Yao Lv ◽  
Chu-di Zhang ◽  
Yi-long Wang ◽  
Dong-ming Zhou ◽  
Meng-ying Zhu ◽  
...  
Keyword(s):  
Side Effects ◽  
Lipid Rafts ◽  
Cell Lines ◽  
Lipid Raft ◽  
Cell Apoptosis ◽  
Cellular Proliferation ◽  
Acid Sphingomyelinase ◽  
Synergistic Cytotoxicity ◽  
Lipid Raft Microdomains

Abstract Background DDP-based chemotherapy is one of the first-line treatment in GC. However, the therapeutic efficacy of DDP is limited due to side effects. Therefore, it is of great significance to develop novel adjuvants to synergize with DDP. We had demonstrated previously that rMV-Hu191 had antitumor activity in GC. Here we examined the synergism of rMV-Hu191 with DDP in vitro and in vivo. Methods Cellular proliferation, the synergistic effect and cell apoptosis were evaluated by CCK-8 assay, ZIP analysis and flow cytometry, respectively. The protein levels and location of ASMase were monitored by western blot and immunofluorescence assay. shRNA and imipramine were used to regulate the expression and activity of ASMase. MβCD was administrated to disrupt lipid rafts. Mice bearing GC xenografts were used to confirm the synergism in vivo. Results From our data, combinational therapy demonstrated synergistic cytotoxicity both in resistant GC cell lines from a Chinese patient and drug-nonresistant GC cell lines, and increased cell apoptosis, instead of viral replication. Integrity of lipid rafts and ASMase were required for rMV-Hu191- and combination-induced apoptosis. The ASMase was delivered to the lipid raft microdomains at the initial stage of rMV-Hu191 treatment. In vivo GC mice xenografts confirmed the synergism of combinational treatment, together with increased apoptosis and trivial side-effects. Conclusions This is the first study to demonstrate that rMV-Hu191 combined with DDP could be used as a potential therapeutic strategy in GC treatment and the ASMase and the integrity of lipid rafts are required for the synergistic effects.

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