A novel kind of tumor type-characteristic junction: plakophilin-2 as a major protein of adherens junctions in cardiac myxomata

Abstract

Using novel antibodies of high avidity to-and specificity for-the constitutive desmosomal plaque protein, plakophilin-2 (Pkp2), in a systematic study of the molecular composition of junctions connecting the cells of soft tissue tumors, we have discovered with immunocytochemical, biochemical and electron microscopical methods, a novel type of adherens junctions in all 32 cardiac myxomata examined. These junctions contain cadherin-11 as their major transmembrane glycoprotein, which we could repeatedly show in colocalization with N-cadherin, anchored in a cytoplasmic plaque formed by a-and b-catenin, together with the further armadillo-type proteins plakoglobin, p120, p0071 and ARVCF. Surprisingly, all adherens junctions of these tumors contained, in addition, another major armadillo protein Pkp2, hitherto known as an obligatory and characteristic constituent of desmosomes in epithelium-derived tumors. We have not detected Pkp2 in a series of noncardiac myxomata studied in parallel. Therefore, we conclude that this acquisition of Pkp2, which we have recently also observed in some mesenchymally derived cells growing in culture, can also occur in tumorigenic transformations in situ. We propose to examine the marker value of Pkp2 in clinical diagnoses of cardiac myxomata and to develop Pkp2-targeted therapeutic reagents.

Intercellular adhering junctions with an asymmetric molecular composition: desmosomes connecting Merkel cells and keratinocytes

“…It seems surprising that Pkp2 has been detected so late as a highly MC-specific marker in the epidermis and its appendages and until very recently its presence in MC-keratinocytes has even been formally denied (e.g., Werling et al 2011). Our discovery of Pkp2 in MCs may partly be the result of a new generation of very sensitive Pkp2-antibodies, as is also suggested by recent findings in certain tumor cells and cell cultures (e.g., Barth et al 2009;Rickelt et al 2009Rickelt et al , 2010.…”

“…ECM extracellular matrix, K keratinocyte, MC Merkel cell. Scale bars: 0.2 μm (a, b) and 0.1 μm (c-e) various proliferatively transformed cell cultures of malignant "soft tissue tumors" (e.g., Barth et al 2009;Rickelt et al 2009Rickelt et al , 2010; for reviews see Franke et al 2009;Pieperhoff et al 2010). It seems surprising that Pkp2 has been detected so late as a highly MC-specific marker in the epidermis and its appendages and until very recently its presence in MC-keratinocytes has even been formally denied (e.g., Werling et al 2011).…”

“…For immunofluorescence and immunoelectron microscopical analyses, the monoclonal as well as the polyclonal antibodies used are listed in Table 1 (see also Franke and Rickelt 2011;Rickelt et al 2010). Primary antibody complexes were visualized with secondary antibodies coupled to Cy3 (Dianova, Hamburg, Germany) or Alexa 488 (MoBiTec, Goettingen, Germany).…”

“…For confocal laser scanning microscopy a Zeiss LSM 510 Meta microscope was used. For immunolocalizations on paraffin-embedded tissue samples, sections of mammalian epidermis were deparaffinized and subjected to antigen retrieval treatment by heat according to standard protocols (see the anthology of Shi et al 2000; for details see Giberson and Demaree 2001;Rickelt et al 2010). Electron as well as immunoelectron microscopy protocols were essentially as described (see, e.g., Langbein et al 2002;Rickelt et al 2008;Schmitt et al 2007).…”

Protein myozap — a late addition to the molecular ensembles of various kinds of adherens junctions

“…Single-and double-label immunofluorescence microscopy was performed as described Rickelt et al 2010), applying, however, increasing numbers of buffer extraction and washing steps or increasing urea concentrations (see also Achtstaetter et al 1986;Franke et al 1983). In the case of paraffin-embedded tissue sections, the antigen-retrieval technique was used (Pieperhoff et al 2011; see also Shi et al 2000).…”