The Desmosome is a Mesoscale Lipid Raft-Like Membrane Domain

Lewis, Joshua E.; Caldara, Amber L; Zimmer, Stephanie; Seybold, Anna; Strong, N.; Stahley, Sara N.; Frangakis, Achilleas S.; Levental, Ilya; Wahl, James K.; Mattheyses, Alexa L.; Sasaki, Takashi; Nakabayashi, Kazuhiko; Hata, Kenichiro; Matsubara, Yoichi; Ishida‐Yamamoto, Akemi; Amagai, Masayuki; Kubo, Akiharu; Kowalczyk, Andrew P.

doi:10.1101/401455

Cited by 14 publications

(17 citation statements)

References 83 publications

(175 reference statements)

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“…Palmitoylation is a covalent attachment of the 16C-fatty acid, palmitate, to cysteine residues through thioester linkage, important for proper protein localisation, association with cellular membranes and lipid rafts, protein trafficking and desmosome assembly [ 20 , 27 – 30 ]. Disruption of palmitoylation has been implicated in several disease states including cancer and treatments that target palmitoylated proteins or palmitoylacyltransferases (PATs) have the potential therapeutic benefits [ 31 ].…”

Section: Resultsmentioning

confidence: 99%

miRNA‐ and cytokine‐associated extracellular vesicles mediate squamous cell carcinomas

Flemming

Hill

Haque

et al. 2020

J of Extracellular Vesicle

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Exosomes, or small extracellular vesicles (sEVs), serve as intercellular messengers with key roles in normal and pathological processes. Our previous work had demonstrated that Dsg2 expression in squamous cell carcinoma (SCC) cells enhanced both sEV secretion and loading of pro-mitogenic cargo. In this study, using wild-type Dsg2 and a mutant form that is unable to be palmitoylated (Dsg2cacs), we investigated the mechanism by which Dsg2 modulates SCC tumour development and progression through sEVs. We demonstrate that palmitoylation was required for Dsg2 to regulate sub-cellular localisation of lipid raft and endosomal proteins necessary for sEV biogenesis. Pharmacological inhibition of the endosomal pathway abrogated Dsg2-mediated sEV release. In murine xenograft models, Dsg2-expressing cells generated larger xenograft tumours as compared to cells expressing GFP or Dsg2cacs. Co-treatment with sEVs derived from Dsg2over-expressing cells increased xenograft size. Cytokine profiling revealed, Dsg2 enhanced both soluble and sEV-associated IL-8 and miRNA profiling revealed, Dsg2 down-regulated both cellular and sEV-loaded miR-146a. miR-146a targets IRAK1, a serine-threonine kinase involved in IL-8 signalling. Treatment with a miR-146a inhibitor up-regulated both IRAK1 and IL-8 expression. RNAseq analysis of HNSCC tumours revealed a correlation between Dsg2 and IL-8. Finally, elevated IL-8 plasma levels were detected in a subset of HNSCC patients who did not respond to immune checkpoint therapy, suggesting that these patients may benefit from prior anti-IL-8 treatment. In summary, these results suggest that intercellular communication through cell-cell adhesion, cytokine release and secretion of EVs are coordinated, and critical for tumour growth and development, and may serve as potential prognostic markers to inform treatment options.

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Section: Resultsmentioning

confidence: 99%

miRNA‐ and cytokine‐associated extracellular vesicles mediate squamous cell carcinomas

Flemming

Hill

Haque

et al. 2020

J of Extracellular Vesicle

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“…These GPMVs fill a unique niche, maintaining the approximate lipid and protein content of living membranes while offering the experimental tractability of synthetic models (3). This combination has allowed investigation of PM composition (4), physical properties (5)(6)(7), dynamics (8,9), and potentially signaling and transport (8,(10)(11)(12)(13) in isolation from the complexity of living cells.…”

Section: Introductionmentioning

confidence: 99%

Cell-derived plasma membrane vesicles are permeable to hydrophilic macromolecules

Skinkle

Levental

2019

Preprint

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Giant Plasma Membrane Vesicles (GPMVs) are a widely used model system for biochemical and biophysical analysis of the isolated mammalian plasma membrane (PM). A core advantage of these vesicles is that they maintain the native lipid and protein diversity of the plasma membrane while affording the experimental flexibility of synthetic giant vesicles. In addition to fundamental investigations of PM structure and composition, GPMVs have been used to evaluate the binding of proteins and small molecules to cell-derived membranes, and the permeation of drug-like molecules through them. An important assumption of such experiments is that GPMVs are sealed; i.e. that permeation occurs by diffusion through the hydrophobic core rather than through hydrophilic pores. Here we demonstrate that this assumption is often incorrect. We find that most GPMVs isolated using standard preparations are passively permeable to various hydrophilic solutes as large as 40 kDa, in contrast to synthetic giant unilamellar vesicles (GUVs). We attribute this leakiness to relatively large and heterogeneous pores formed by rupture of vesicles from cells. These pores are stable and persist throughout experimentally relevant time scales. Finally, we identify preparation conditions that minimize poration and allow evaluation of sealed GPMVs. These unexpected observations of GPMV poration are of critical importance for interpreting experiments utilizing GPMVs as plasma membrane models, particularly for drug permeation and membrane asymmetry. STATEMENT OF SIGNIFICANCE:A critical assumption in using Giant Plasma Membrane Vesicles to study membrane penetration and interactions is that these vesicles maintain the permeability barrier of the native membrane from which they form. Using large fluorescently-labeled hydrophilic probes, we demonstrate that this assumption is often incorrect and conclude that macromolecular solutes permeate GPMVs through stable pores formed during shear-induced rupture of vesicles from cells. Using these insights into the mechanisms of poration, we demonstrate an approach to isolate sealed GPMVs.

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“…Adherens junction and tight junction molecules are essential for the maintenance of the laryngeal epithelium structure, contributing to the protection and functional activity of this tissue [ 78 ]. Moreover, desmosomes are crucial for the integrity of tissues that undergo constant mechanical stress as do vocal folds [ 79 , 80 ]. Interestingly, the literature reports alterations in the laryngeal epithelial barrier due to phonotrauma and LPR.…”

Section: Discussionmentioning

confidence: 99%

Unraveling the molecular pathobiology of vocal fold systemic dehydration using an in vivo rabbit model

et al. 2020

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Vocal folds are a viscoelastic multilayered structure responsible for voice production. Vocal fold epithelial damage may weaken the protection of deeper layers of lamina propria and thyroarytenoid muscle and impair voice production. Systemic dehydration can adversely affect vocal function by creating suboptimal biomechanical conditions for vocal fold vibration. However, the molecular pathobiology of systemically dehydrated vocal folds is poorly understood. We used an in vivo rabbit model to investigate the complete gene expression profile of systemically dehydrated vocal folds. The RNA-Seq based transcriptome revealed 203 differentially expressed (DE) vocal fold genes due to systemic dehydration. Interestingly, function enrichment analysis showed downregulation of genes involved in cell adhesion, cell junction, inflammation, and upregulation of genes involved in cell proliferation. RT-qPCR validation was performed for a subset of DE genes and confirmed the downregulation of DSG1 , CDH3 , NECTIN1 , SDC1 , S100A9 , SPINK5 , ECM1 , IL1A , and IL36A genes. In addition, the upregulation of the transcription factor NR4A3 gene involved in epithelial cell proliferation was validated. Taken together, these results suggest an alteration of the vocal fold epithelial barrier independent of inflammation, which could indicate a disruption and remodeling of the epithelial barrier integrity. This transcriptome provides a first global picture of the molecular changes in vocal fold tissue in response to systemic dehydration. The alterations observed at the transcriptional level help to understand the pathobiology of dehydration in voice function and highlight the benefits of hydration in voice therapy.

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The Desmosome is a Mesoscale Lipid Raft-Like Membrane Domain

Cited by 14 publications

References 83 publications

miRNA‐ and cytokine‐associated extracellular vesicles mediate squamous cell carcinomas

miRNA‐ and cytokine‐associated extracellular vesicles mediate squamous cell carcinomas

Cell-derived plasma membrane vesicles are permeable to hydrophilic macromolecules

Unraveling the molecular pathobiology of vocal fold systemic dehydration using an in vivo rabbit model

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