Plakophilin 1 but not plakophilin 3 regulates desmoglein clustering

  • Michael Fuchs
  • Marco Foresti
  • Mariya Y. Radeva
  • Daniela Kugelmann
  • Rene Keil
  • Mechthild Hatzfeld
  • Volker Spindler
  • Jens WaschkeEmail author
  • Franziska VielmuthEmail author
Original Article


Plakophilins (Pkp) are desmosomal plaque proteins crucial for desmosomal adhesion and participate in the regulation of desmosomal turnover and signaling. However, direct evidence that Pkps regulate clustering and molecular binding properties of desmosomal cadherins is missing. Here, keratinocytes lacking either Pkp1 or 3 in comparison to wild type (wt) keratinocytes were characterized with regard to their desmoglein (Dsg) 1- and 3-binding properties and their capability to induce Dsg3 clustering. As revealed by atomic force microscopy (AFM), both Pkp-deficient keratinocyte cell lines showed reduced membrane availability and binding frequency of Dsg1 and 3 at cell borders. Extracellular crosslinking and AFM cluster mapping demonstrated that Pkp1 but not Pkp3 is required for Dsg3 clustering. Accordingly, Dsg3 overexpression reconstituted cluster formation in Pkp3- but not Pkp1-deficient keratinocytes as shown by AFM and STED experiments. Taken together, these data demonstrate that both Pkp1 and 3 regulate Dsg membrane availability, whereas Pkp1 but not Pkp3 is required for Dsg3 clustering.


Desmosome Cell adhesion Desmosomal clustering Atomic force microscopy STED 



Atomic force microscopy


Arrhythmogenic cardiomyopathy








Extracellular domains


Fluorescence recovery after photobleaching


Cultured human keratinocytes


Intermediate filament


Mitogen-activated protein kinase


Murine keratinocytes








Pemphigus vulgaris


Quantitative imaging


Stimulated emission depletion


Ethylene glycolbis (sulfosuccinimidylsuccinate)


Unbinding forces


Unbinding position


Wild type



We thank Andrea Wehmeyer and Sabine Mühlsimer for excellent technical assistance and JPK instruments and Sunil Yeruva for constructive technical and scientific discussion. The project is funded by Else-Kröner-Fresenius-Stiftung 2016_AW157 to FV and JW and DFG SPP1782 to JW.

Author contributions

MFu, MF, MR, DK: methodology, data acquisition and analysis; FV and JW: funding acquisition, conceptualization and supervision; MFu, FV, JW: writing and editing, RK, MH, VS: methodology, review.

Compliance with ethical standards

Conflict of interest

The authors state that there was no conflict of interest.

Supplementary material

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Supplementary material 1 (TIFF 25518 kb)
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Supplementary material 2 (TIFF 25518 kb)
18_2019_3083_MOESM3_ESM.tif (24.9 mb)
Supplementary material 3 (TIFF 25518 kb)
18_2019_3083_MOESM4_ESM.docx (25 kb)
Supplementary material 4 (DOCX 24 kb)


  1. 1.
    Delva E, Tucker KD, Kowalczyk AP (2009) The desmosome. Cold Spring Harb Perspect Biol 1:a002543Google Scholar
  2. 2.
    Spindler V, Waschke J (2014) Desmosomal cadherins and signaling: lessons from autoimmune disease. Cell Commun Adhes 21:8Google Scholar
  3. 3.
    Delmar M, McKenna WJ (2010) The cardiac desmosome and arrhythmogenic cardiomyopathies from gene to disease. Circ Res 107:15Google Scholar
  4. 4.
    McGrath JA, Mellerio JE (2010) Ectodermal dysplasia-skin fragility syndrome. Dermatol Clin 28(1):125–129Google Scholar
  5. 5.
    Vielmuth F, Spindler V, Waschke J (2018) Atomic force microscopy provides new mechanistic insights into the pathogenesis of pemphigus. Front Immunol 9:8Google Scholar
  6. 6.
    Harrison OJ et al (2016) Structural basis of adhesive binding by desmocollins and desmogleins. Proc Natl Acad Sci USA 113(26):6Google Scholar
  7. 7.
    Nie Z et al (2011) Membrane-impermeable Cross-linking provides evidence for homophilic, isoform-specific binding of desmosomal cadherins in epithelial cells. J Biol Chem 286(3):11Google Scholar
  8. 8.
    Hobbs R, Green K (2011) Desmoplakin regulates desmosome hyperadhesion. J Investig Dermatol 132:482Google Scholar
  9. 9.
    Waschke J (2008) The desmosome and pemphigus. Histochem Cell Biol 130:21–54Google Scholar
  10. 10.
    Waschke J, Spindler V (2014) Desmosomes and extradesmosomal adhesive signaling contacts in pemphigus. Med Res Rev 34:1127–1145Google Scholar
  11. 11.
    Bass-Zubeck AE et al (2009) Plakophilins: multifunctional scaffolds for adhesion and signaling. Curr Opin Cell Biol 21:708–716Google Scholar
  12. 12.
    Nekrasova O, Green KJ (2013) Desmosome assembly and dynamics. Trends Cell Biol 23(11):10Google Scholar
  13. 13.
    Hatzfeld M, Wolf A, Keil R (2014) Plakophilins in desmosomal adhesion and signaling. Cell Commun Adhes 21:708–716Google Scholar
  14. 14.
    Keil R, Rietscher K, Hatzfeld M (2016) Antagonistic regulation of intercellular cohesion by plakophilins 1 and 3. J Invest Dermatol 136(10):2022–2029. Google Scholar
  15. 15.
    McGrath JA et al (1997) Mutations in the plakophilin 1 gene result in ectodermal dysplasia/skin fragility syndrome. Nat Genet 17(2):240–244Google Scholar
  16. 16.
    Boyce AE et al (2012) Ectodermal dysplasia-skin fragility syndrome due to a new homozygous internal deletion mutation in the PKP1 gene. Aust J Dermatol 53(1):61–65Google Scholar
  17. 17.
    Rietscher K et al (2016) Growth retardation, loss of desmosomal adhesion and impaired tight junction function identify a unique role of plakophilin 1 in vivo. J Invest Dermatol 136(7):1471–1478. Google Scholar
  18. 18.
    Sklyarova T et al (2008) Plakophilin-3-deficient mice develop hair coat abnormalities and are prone to cutaneous inflammation. J Invest Dermatol 128(6):1375–1385Google Scholar
  19. 19.
    Al-Amoudi A et al (2011) The three-dimensional molecular structure of the desmosomal plaque. Proc Natl Acad Sci USA 108(16):6Google Scholar
  20. 20.
    Kowalczyk AP et al (1999) The head domain of plakophilin-1 binds to desmoplakin and enhances its recruitment to desmosomes: implications for cutaneous disease. J Biol Chem 274(26):5Google Scholar
  21. 21.
    Bonné S et al (2003) Defining desmosomal plakophilin-3 interactions. J Cell Biol 161(2):14Google Scholar
  22. 22.
    Hatzfeld M et al (2000) The function of plakophilin 1 in desmosome assembly and actin filament organization. J Cell Biol 149(1):209–222Google Scholar
  23. 23.
    Fischer-Kešo R et al (2014) Plakophilins 1 and 3 bind to FXR1 and thereby influence the mRNA stability of desmosomal proteins. Mol Cell Biol 34(23):13Google Scholar
  24. 24.
    Lee P et al (2017) Phosphorylation of Pkp1 by RIPK4 regulates epidermal differentiation and skin tumorigenesis. EMBO J 13:1963–1980Google Scholar
  25. 25.
    Neuber S et al (2010) The desmosomal plaque proteins of the plakophilin family. Dermatol Res Pract 2010:11Google Scholar
  26. 26.
    Tucker KD, Stahley S, Kowalczyk AP (2014) Plakophilin-1 protects keratinocytes from pemphigus vulgaris IgG by forming calcium-independent desmosomes. J Investig Dermatol 134:11Google Scholar
  27. 27.
    Garrod D, Tabernero L (2014) Hyper-adhesion: a unique property of desmosomes. Cell Commun Adhes. 21:8Google Scholar
  28. 28.
    Stahley SN et al (2016) Super-resolution microscopy reveals altered desmosomal protein organization in tissue from patients with pemphigus vulgaris. J Invest Dermatol 136(1):59–66Google Scholar
  29. 29.
    Winik BC et al (2009) Acantholytic ectodermal dysplasia: clinicopathological study of a new desmosomal disorder. Br J Dermatol 160(4):868–874Google Scholar
  30. 30.
    Diercks GF, Pas HH, Jonkman MF (2009) The ultrastructure of acantholysis in pemphigus vulgaris. Br J Dermatol 160(2):460–461Google Scholar
  31. 31.
    Oktarina DA et al (2011) IgG-induced clustering of desmogleins 1 and 3 in skin of patients with pemphigus fits with the desmoglein nonassembly depletion hypothesis. Br J Dermatol 165(3):552–562Google Scholar
  32. 32.
    van der Wier G et al (2014) Smaller desmosomes are seen in the skin of pemphigus patients with anti-desmoglein 1 antibodies but not in patients with anti-desmoglein 3 antibodies. J Invest Dermatol 134(8):2287–2290Google Scholar
  33. 33.
    Sokol E et al (2015) Large-scale electron microscopy maps of patient skin and mucosa provide insight into pathogenesis of blistering diseases. J Invest Dermatol 135(7):1763–1770Google Scholar
  34. 34.
    Vielmuth F et al (2018) Keratins regulate the adhesive properties of desmosomal cadherins through signaling. J Investig Dermatol 138:11Google Scholar
  35. 35.
    Spindler V et al (2018) Mechanisms causing loss of keratinocyte cohesion in pemphigus. J Invest Dermatol 138(1):6Google Scholar
  36. 36.
    Kasperkiewicz M et al (2017) Pemphigus. Nat Rev Dis Prim 3:17026Google Scholar
  37. 37.
    Boukamp P et al (1988) Normal keratinization in a spontaneously immortalized aneuploid human keratinocyte cell line. J Cell Biol 106(3):11Google Scholar
  38. 38.
    Heupel WM et al (2008) Pemphigus vulgaris IgG directly inhibit desmoglein 3-mediated transinteraction. J Immunol 181(3):10Google Scholar
  39. 39.
    Waschke J et al (2005) Pemphigus foliaceus IgG causes dissociation of desmoglein 1-containing junctions without blocking desmoglein 1 transinteraction. J Clin Invest 115(11):3157–3165Google Scholar
  40. 40.
    Vielmuth F et al (2015) Atomic force microscopy identifies regions of distinct desmoglein 3 adhesive properties on living keratinocytes. Nanomed Nanotechnol Biol Med 11(3):10Google Scholar
  41. 41.
    Vielmuth F, Waschke J, Spindler V (2015) Loss of desmoglein binding is not sufficient for keratinocyte dissociation in pemphigus. J Investig Dermatol 135(12):10Google Scholar
  42. 42.
    Ebner A et al (2007) A new, simple method for linking of antibodies to atomic force microscopy tips. Bioconj Chem 18(4):9Google Scholar
  43. 43.
    Hartlieb E et al (2013) Desmoglein 2 is less important than desmoglein 3 for keratinocyte cohesion. PLoS One 8(1):12Google Scholar
  44. 44.
    Spindler V et al (2013) Peptide-mediated desmoglein 3 crosslinking prevents pemphigus vulgaris autoantibody-induced skin blistering. J Clin Investig 123(2):12Google Scholar
  45. 45.
    Kowalczyk AP, Green KJ (2015) Structure, function and regulation of desmosomes. Prog Mol Biol Transl 116:23Google Scholar
  46. 46.
    Waugh R, Hochmuth R (1987) Mechanical equilibrium of thick, hollow, liquid membrane cylinders. Biophys J 52(3):10Google Scholar
  47. 47.
    Schmitz J, Benoit M, Gottschalk KE (2008) The viscoelasticity of membrane tethers and its importance for cell adhesion. Biophys J 95:12Google Scholar
  48. 48.
    Sariisik E et al (2015) Decoding cytoskeleton-anchored and non-anchored receptors from single-cell adhesion force data. Biophys J 109(7):4Google Scholar
  49. 49.
    Stanley JR, Amagai M (2006) Pemphigus, bullous impetigo, and the staphylococcal scalded-skin syndrome. N Engl J Med 355(17):10Google Scholar
  50. 50.
    Spindler V, Waschke J (2018) Pemphigus—a disease of desmosome dysfunction caused by multiple mechanisms. Front Immunol 9:136Google Scholar
  51. 51.
    Vielmuth F et al (2018) Keratins regulate p38MAPK-dependent desmoglein binding properties in pemphigus. Front Immunol. 9:528Google Scholar
  52. 52.
    Berkowitz P et al (2006) p38MAPK inhibition prevents disease in pemphigus vulgaris mice. Proc Natl Acad Sci USA 103:34Google Scholar
  53. 53.
    Sariisik E et al (2015) Decoding cytoskeleton-anchored and non-anchored receptors from single-cell adhesion force data. Biophys J 109(7):1330–1333Google Scholar
  54. 54.
    Hatzfeld M (2007) Plakophilins: Multifunctional proteins or just regulators of desmosomal adhesion? BBA Mol Cell Res 1773(1):9Google Scholar
  55. 55.
    Basu S et al (2018) Plakophilin3 loss leads to an increase in lipocalin2 expression, which is required for tumour formation. Exp Cell Res. 369:251–265Google Scholar
  56. 56.
    Muller DJ (2008) AFM: a nanotool in membrane biology. Biochemistry 47:13Google Scholar
  57. 57.
    Zlatanova J, Lindsya SM, Sanford HL (2000) Single molecule force spectroscopy in biology using the atomic force microscope. Prog Biophys Mol Biol 74:25Google Scholar
  58. 58.
    Cirillo N et al (2006) Serum from pemphigus vulgaris reduces desmoglein 3 half-life and perturbs its de novo assembly to desmosomal sites in cultured keratinocytes. FEBS Lett 580(13):3276–3281Google Scholar
  59. 59.
    South AP et al (2003) Lack of plakophilin 1 increases keratinocyte migration and reduces desmosome stability. J Cell Sci 116:12Google Scholar
  60. 60.
    Gurjar M et al (2017) Plakophilin3 increases desmosome assembly, size and stability by increasing expression of desmocollin2. Biochem Biophys Res Commun 495:768–774Google Scholar
  61. 61.
    Todorovic V et al (2014) Plakophilin 3 mediates Rap1-dependent desmosome assembly and adherens junction maturation. Mol Biol Cell 25(23):15Google Scholar
  62. 62.
    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):1033–1043Google Scholar
  63. 63.
    Egu DT et al (2017) Inhibition of p38MAPK signalling prevents epidermal blistering and alterations of desmosome structure induced by pemphigus autoantibodies in human epidermis. Br J Dermatol. 177(6):1612–1618Google Scholar
  64. 64.
    Spindler V et al (2014) Plakoglobin but not desmoplakin regulates keratinocyte cohesion via modulation of p38MAPK signaling. J Invest Dermatol 134(6):10Google Scholar
  65. 65.
    Kusumi A, Suzuki K, Koyasako K (1999) Mobility and cytoskeletal interactions of cell adhesion receptors. Curr Opin Cell Biol 11(5):8Google Scholar
  66. 66.
    Truong Quang BA et al (2013) Principles of E-cadherin supramolecular organization in vivo. Curr Biol 23(22):11Google Scholar
  67. 67.
    Wu Y, Kanchanawong P, Zaidel-Bar R (2015) Actin-delimited adhesion-independent clustering of E-cadherin forms the nanoscale building blocks of adherens junctions. Dev Cell 32(2):6Google Scholar
  68. 68.
    Owen GR, Stokes DL (2010) Exploring the nature of desmosomal cadherin associations in 3D. Dermatol Res Pract 2010:930401Google Scholar
  69. 69.
    Stahley SN et al (2016) Molecular organization of the desmosome as revealed by direct stochastic optical reconstruction microscopy. J Cell Sci 129(15):2897–2904Google Scholar

Copyright information

© Springer Nature Switzerland AG 2019

Authors and Affiliations

  • Michael Fuchs
    • 1
  • Marco Foresti
    • 1
  • Mariya Y. Radeva
    • 1
  • Daniela Kugelmann
    • 1
  • Rene Keil
    • 2
  • Mechthild Hatzfeld
    • 2
  • Volker Spindler
    • 3
  • Jens Waschke
    • 1
    Email author
  • Franziska Vielmuth
    • 1
    Email author
  1. 1.Faculty of Medicine, Institute of AnatomyLudwig-Maximilians-Universität MunichMunichGermany
  2. 2.Division of Pathobiochemistry, Institute of Molecular MedicineMartin-Luther-University Halle-WittenbergHalleGermany
  3. 3.Department of BiomedicineUniversity of BaselBaselSwitzerland

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