Histochemistry and Cell Biology

, Volume 152, Issue 3, pp 195–206 | Cite as

Depletion of the cellular cholesterol content reduces the dynamics of desmosomal cadherins and interferes with desmosomal strength

  • Nataša Resnik
  • Giulia Maria Rita de Luca
  • Kristina Sepčić
  • Rok Romih
  • Erik Manders
  • Peter VeraničEmail author
Original Paper


Desmosomal cadherins, desmocollins, and desmogleins are cholesterol-dependent entities responsible for the stable adhesion of desmosomes in epithelial cells. Here, we investigated the influence of cellular cholesterol depletion on the dynamic properties of the desmosomal cadherin desmocollin, particularly the lateral mobility and distribution of desmocollin 2 (Dsc2-YFP) in the plasma membrane, and how these properties influence the adhesion strength of desmosomes. Depletion of cellular cholesterol decreased the lateral mobility of Dsc2-YFP and caused dispersion of Dsc2-YFP in the plasma membrane of epithelial MDCK cells. As a consequence of the altered Dsc2-YFP dynamics, the adhesive strength of desmosomes was weakened. Moreover, our study is the first to show and quantify the co-association of desmosomes with cholesterol/sphingomyelin-enriched membrane domains at the ultrastructural level. Taken together, our data emphasize a critical role for the cellular cholesterol content in regulating the lateral mobility and distribution of Dsc2 and show that cholesterol depletion reduces the strength of desmosomal adhesions.


Cholesterol Desmosomes Desmosomal cadherins Lateral mobility Cholesterol/sphingomyelin-enriched membrane domains 



Bovine serum albumin


Diffusion coefficient


Desmocollin 2 tagged with yellow fluorescent protein


Fluorescence recovery after photobleaching




Mobile fraction


Ostreolysin A/pleurotolysin B


Phosphate-buffered saline



The study was supported by Slovenian Research Agency (ARRS) grants P3-0108, P1-0207 and J3-7494 and MRIC UL IP-0510 Infrastructure program. MDCK cells with Dsc2-YFP expression were kindly provided by Prof. Rudolf Leube (University Hospital RWTH Aachen, Germany). We express gratitude to Ronald Breedijk, Sabina Železnik, Sanja Čabraja, Linda Štrus, and Nada Pavlica Dubarič for their technical assistance.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Supplementary material

418_2019_1797_MOESM1_ESM.docx (225 kb)
Supplementary material 1 (DOCX 224 kb)


  1. Baier CJ, Gallegos CE, Levi V, Barrantes FJ (2010) Cholesterol modulation of nicotinic acetylcholine receptor surface mobility. Eur Biophys J 39(2):213–227. CrossRefPubMedGoogle Scholar
  2. Bashour KT, Tsai J, Shen K, Lee JH, Sun E, Milone MC, Dustin ML, Kam LC (2014) Cross talk between CD3 and CD28 is spatially modulated by protein lateral mobility. Mol Cell Biol 34(6):955–964. CrossRefPubMedPubMedCentralGoogle Scholar
  3. Berne S, Križaj I, Pohleven F, Turk T, Maček P, Sepčić K (2002) Pleurotus and Agrocybe hemolysins, new proteins hypothetically involved in fungal fruiting. Biochim Biophys Acta 1570(3):153–159CrossRefPubMedGoogle Scholar
  4. Bligh EG, Dyer WJ (1959) A rapid method of total lipid extraction and purification. Can J Biochem Physiol 37(8):911–917. CrossRefPubMedGoogle Scholar
  5. Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248–254CrossRefPubMedPubMedCentralGoogle Scholar
  6. Causeret M, Taulet N, Comunale F, Favard C, Gauthier-Rouvière C (2005) N-cadherin association with lipid rafts regulates its dynamic assembly at cell-cell junctions in C2C12 myoblasts. Mol Biol Cell 16(5):2168–2180. CrossRefPubMedPubMedCentralGoogle Scholar
  7. De Luca GMR, Desclos E, Breedijk RMP, Dolz-Edo L, Smits GJ, Nahidiazar L, Bielefeld P, Picavet L, Fitzsimons CP, Hoebe R, Manders EMM (2017) Configurations of the Re-scan confocal microscope (RCM) for biomedical applications. J Microsc 266(2):166–177. CrossRefGoogle Scholar
  8. Gloushankova NA, Wakatsuki T, Troyanovsky RB, Elson E, Troyanovsky SM (2003) Continual assembly of desmosomes within stable intercellular contacts of epithelial A-431 cells. Cell Tissue Res 314(3):399–410. CrossRefPubMedGoogle Scholar
  9. Hiramoto-Yamaki N, Tanaka KA, Suzuki KG, Hirosawa KM, Miyahara MS, Kalay Z, Tanaka K, Kasai RS, Kusumi A, Fujiwara TK (2014) Ultrafast diffusion of a fluorescent cholesterol analog in compartmentalized plasma membranes. Traffic 15(6):583–612. CrossRefPubMedPubMedCentralGoogle Scholar
  10. Huen AC, Park JK, Godsel LM, Chen X, Bannon LJ, Amargo EV, Hudson TY, Mongiu AK, Leigh IM, Kelsell DP, Gumbiner BM, Green KJ (2002) Intermediate filament-membrane attachments function synergistically with actin-dependent contacts to regulate intercellular adhesive strength. J Cell Biol 159(6):1005–1017. CrossRefPubMedPubMedCentralGoogle Scholar
  11. Ihrie RA, Attardi LD (2005) A new Perp in the lineup: linking p63 and desmosomal adhesion. Cell Cycle 4(7):873–876. CrossRefPubMedGoogle Scholar
  12. Iino R, Koyama I, Kusumi A (2001) Single molecule imaging of green fluorescent proteins in living cells: E-cadherin forms oligomers on the free cell surface. Biophys J 80(6):2667–2677. CrossRefPubMedPubMedCentralGoogle Scholar
  13. Kenworthy AK, Nichols BJ, Remmert CL, Hendrix GM, Kumar M, Zimmerberg J, Lippincott-Schwartz J (2004) Dynamics of putative raft-associated proteins at the cell surface. J Cell Biol 165(5):735–746. CrossRefPubMedPubMedCentralGoogle Scholar
  14. Klonis N, Rug M, Harper I, Wickham M, Cowman A, Tilley L (2002) Fluorescence photobleaching analysis for the study of cellular dynamics. Eur Biophys J 31(1):36–51CrossRefPubMedGoogle Scholar
  15. Kowalczyk AP, Green KJ (2013) Structure, function, and regulation of desmosomes. Prog Mol Biol Transl Sci 116:95–118. CrossRefPubMedPubMedCentralGoogle Scholar
  16. Lang T, Bruns D, Wenzel D, Riedel D, Holroyd P, Thiele C, Jahn R (2001) SNAREs are concentrated in cholesterol-dependent clusters that define docking and fusion sites for exocytosis. EMBO J 20(9):2202–2213. CrossRefPubMedPubMedCentralGoogle Scholar
  17. Lingwood D, Simons K (2010) Lipid rafts as a membrane-organizing principle. Science 327(5961):46–50. CrossRefGoogle Scholar
  18. Nekrasova O, Green KJ (2013) Desmosome assembly and dynamics. Trends Cell Biol 23(11):537–546. CrossRefPubMedPubMedCentralGoogle Scholar
  19. Nishimura SY, Vrljic M, Klein LO, McConnell HM, Moerner WE (2006) Cholesterol depletion induces solid-like regions in the plasma membrane. Biophys J 90(3):927–938. CrossRefPubMedGoogle Scholar
  20. Ohvo H, Slotte JP (1996) Cyclodextrin-mediated removal of sterols from monolayers: effects of sterol structure and phospholipids on desorption rate. Biochemistry 35(24):8018–8024. CrossRefPubMedGoogle Scholar
  21. Osmani N, Labouesse M (2015) Remodeling of keratin-coupled cell adhesion complexes. Curr Opin Cell Biol 32:30–38. CrossRefPubMedGoogle Scholar
  22. Ota K, Leonardi A, Mikelj M, Skočaj M, Wohlschlager T, Künzler M, Aebi M, Narat M, Križaj I, Anderluh G, Sepčić K, Maček P (2013) Membrane cholesterol and sphingomyelin, and ostreolysinA are obligatory for pore-formation by a MACPF/CDC-like pore-forming protein, pleurotolysinB. Biochimie 95(10):1855–1864. CrossRefPubMedGoogle Scholar
  23. Pasdar M, Nelson WJ (1988) Kinetics of desmosome assembly in Madin-Darby canine kidney epithelial cells: temporal and spatial regulation of desmoplakin organization and stabilization upon cell–cell contact I. Biochemical analysis. J Cell Biol 106(3):677–685CrossRefPubMedGoogle Scholar
  24. Pasdar M, Krzeminski KA, Nelson WJ (1991) Regulation of desmosome assembly in MDCK epithelial cells: coordination of membrane core and cytoplasmic plaque domain assembly at the plasma membrane. J Cell Biol 113(3):645–655CrossRefPubMedGoogle Scholar
  25. Phair RD, Gorski SA, Misteli T (2004) Measurement of dynamic protein binding to chromatin in vivo, using photobleaching microscopy. Methods Enzymol 375:393–414CrossRefPubMedGoogle Scholar
  26. Resnik N, Sepčić K, Plemenitaš A, Windoffer R, Leube R, Veranič P (2011) Desmosome assembly and cell–cell adhesion are membrane raft-dependent processes. J Biol Chem 286(2):1499–1507. CrossRefPubMedGoogle Scholar
  27. Resnik N, Repnik U, Kreft ME, Sepčić K, Maček P, Turk B, Veranič P (2015) Highly selective anti-cancer activity of cholesterol-interacting agents methyl-β-cyclodextrin and ostreolysinA/pleurotolysinB protein complex on urothelial cancer cells. PLoS One 10(9):e0137878. CrossRefPubMedPubMedCentralGoogle Scholar
  28. Roberts BJ, Johnson KE, McGuinn KP, Saowapa J, Svoboda RA, Mahoney MG, Johnson KR, Wahl JK (2014) Palmitoylation of plakophilin is required for desmosome assembly. J Cell Sci 127(Pt 17):3782–3793. CrossRefPubMedPubMedCentralGoogle Scholar
  29. Roberts BJ, Svoboda RA, Overmiller AM, Lewis JD, Kowalczyk AP, Mahoney MG, Johnson KR, Wahl JK (2016) Palmitoylation of desmoglein 2 is a regulator of assembly dynamics and protein turnover. J Biol Chem 291(48):24857–24865. CrossRefPubMedPubMedCentralGoogle Scholar
  30. Sezgin E, Levental I, Mayor S, Eggeling C (2017) The mystery of membrane organization: composition, regulation and roles of lipid rafts. Nat Rev Mol Cell Biol 18(6):361–374. CrossRefPubMedPubMedCentralGoogle Scholar
  31. Skočaj M, Resnik N, Grundner M, Ota K, Rojko N, Hodnik V, Anderluh G, Sobota A, Maček P, Veranič P, Sepčić K (2014) Tracking cholesterol/sphingomyelin-rich membrane domains with the ostreolysinA-mCherry protein. PLoS One 9(3):e92783. CrossRefPubMedPubMedCentralGoogle Scholar
  32. Stahley SN, Saito M, Faundez V, Koval M, Mattheyses AL, Kowalczyk AP (2014) Desmosome assembly and disassembly are membrane raft-dependent. PLoS One 9(1):e87809. CrossRefPubMedPubMedCentralGoogle Scholar
  33. Tucker DK, Stahley SN, Kowalczyk AP (2014) Plakophilin-1 protects keratinocytes from pemphigus vulgaris IgG by forming calcium-independent desmosomes. J Invest Dermatol 134(4):1033–1043. CrossRefPubMedGoogle Scholar
  34. Völlner F, Ali J, Kurrle N, Exner Y, Eming R, Hertl M, Banning A, Tikkanen R (2016) Loss of flotillin expression results in weakened desmosomal adhesion and Pemphigus vulgaris-like localisation of desmoglein-3 in human keratinocytes. Sci Rep 6:28820. CrossRefPubMedPubMedCentralGoogle Scholar
  35. Windoffer R, Borchert-Stuhlträger M, Leube RE (2002) Desmosomes: interconnected calcium-dependent structures of remarkable stability with significant integral membrane protein turnover. J Cell Sci 115(Pt 8):1717–1732PubMedGoogle Scholar
  36. Zidovetzki R, Levitan I (2007) Use of cyclodextrins to manipulate plasma membrane cholesterol content: evidence, misconceptions and control strategies. Biochim Biophys Acta 1768(6):1311–1324. CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  • Nataša Resnik
    • 1
  • Giulia Maria Rita de Luca
    • 2
  • Kristina Sepčić
    • 3
  • Rok Romih
    • 1
  • Erik Manders
    • 2
  • Peter Veranič
    • 1
    Email author
  1. 1.Faculty of Medicine, Institute of Cell BiologyUniversity of LjubljanaLjubljanaSlovenia
  2. 2.Faculty of Science, Swammerdam Institute for Life Sciences, Leeuwenhoek Centre for Advanced MicroscopyUniversity of AmsterdamAmsterdamThe Netherlands
  3. 3.Department of Biology, Biotechnical FacultyUniversity of LjubljanaLjubljanaSlovenia

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