Reversible Dissolution of Microdomains in Detergent-Resistant Membranes at Physiological Temperature

Cremona, Andrea; Orsini, F.; Corsetto, Paola Antonia; Hoogenboom, Bart W.; Rizzo, Angela Maria

doi:10.1371/journal.pone.0132696

Cited by 2 publications

(1 citation statement)

References 45 publications

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“…Altogether, these data suggest that MβCD-resistant cholesterol in membranes of the MDA-MB-435 cells reflects their richness in lipid rafts. This will be evaluated in a future experiment by atomic force microscopy, as recently proposed ( Cremona et al, 2015 ).…”

Section: Discussionmentioning

confidence: 99%

Cellular Cholesterol Distribution Influences Proteolytic Release of the LRP-1 Ectodomain

et al. 2016

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Low-density lipoprotein receptor-related protein-1 (LRP-1) is a multifunctional matricellular receptor composed of a large ligand-binding subunit (515-kDa α-chain) associated with a short trans-membrane subunit (85-kDa β-chain). LRP-1, which exhibits both endocytosis and cell signaling properties, plays a key role in tumor invasion by regulating the activity of proteinases such as matrix metalloproteinases (MMPs). LRP-1 is shed at the cell surface by proteinases such as membrane-type 1 MMP (MT1-MMP) and a disintegrin and metalloproteinase-12 (ADAM-12). Here, we show by using biophysical, biochemical, and cellular imaging approaches that efficient extraction of cell cholesterol and increased LRP-1 shedding occur in MDA-MB-231 breast cancer cells but not in MDA-MB-435 cells. Our data show that cholesterol is differently distributed in both cell lines; predominantly intracellularly for MDA-MB-231 cells and at the plasma membrane for MDA-MB-435 cells. This study highlights the relationship between the rate and cellular distribution of cholesterol and its impact on LRP-1 shedding modulation. Altogether, our data strongly suggest that the increase of LRP-1 shedding upon cholesterol depletion induces a higher accessibility of the sheddase substrate, i.e., LRP-1, at the cell surface rather than an increase of expression of the enzyme.

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Section: Discussionmentioning

confidence: 99%

Cellular Cholesterol Distribution Influences Proteolytic Release of the LRP-1 Ectodomain

et al. 2016

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Comparative lipidomics and proteomics analysis of platelet lipid rafts using different detergents

et al. 2016

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Lipid rafts play a pivotal role in physiological functions of platelets. Their isolation using nonionic mild detergents is considered as the gold standard method, but there is no consensual detergent for lipid raft studies. We aimed to investigate which detergent is the most suitable for lipid raft isolation from platelet membrane, based on lipidomics and proteomics analysis. Platelets were obtained from healthy donors. Twelve sucrose fractions were extracted by three different detergents, namely Brij 35, Lubrol WX, and Triton X100, at 0.05% and 1%. After lipidomics analysis and determination of fractions enriched in cholesterol (Ch) and sphingomyelin (SM), proteomics analysis was performed. Lipid rafts were mainly observed in 1-4 fractions, and non-rafts were distributed on 5-12 fractions. Considering the concentration of Ch and SM, Lubrol WX 1% and Triton X100 1% were more suitable detergents as they were able to isolate lipid raft fractions that were more enriched than non-raft fractions. By proteomics analysis, overall, 822 proteins were identified in platelet membrane. Lipid raft fractions isolated with Lubrol WX 0.05% and Triton X100 1% contained mainly plasma membrane proteins. However, only Lubrol WX 0.05 and 1% and Triton X100 1% were able to extract non-denaturing proteins with more than 10 transmembrane domains. Our results suggest that Triton X100 1% is the most suitable detergent for global lipid and protein studies on platelet plasma membrane. However, the detergent should be adapted if investigation of an association between specific proteins and lipid rafts is planned.

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Reversible Dissolution of Microdomains in Detergent-Resistant Membranes at Physiological Temperature

Cited by 2 publications

References 45 publications

Cellular Cholesterol Distribution Influences Proteolytic Release of the LRP-1 Ectodomain

Cellular Cholesterol Distribution Influences Proteolytic Release of the LRP-1 Ectodomain

Comparative lipidomics and proteomics analysis of platelet lipid rafts using different detergents

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