The Function of Plakophilin 1 in Desmosome Assembly and Actin Filament Organization

Molecular Biology Group of the Medical Faculty, University of Halle, 06097 Halle/Saale, Germany.
The Journal of Cell Biology (Impact Factor: 9.83). 05/2000; 149(1):209-22.
Source: PubMed

ABSTRACT Plakophilin 1, a member of the armadillo multigene family, is a protein with dual localization in the nucleus and in desmosomes. To elucidate its role in desmosome assembly and regulation, we have analyzed its localization and binding partners in vivo. When overexpressed in HaCaT keratinocytes, plakophilin 1 localized to the nucleus and to desmosomes, and dramatically enhanced the recruitment of desmosomal proteins to the plasma membrane. This effect was mediated by plakophilin 1's head domain, which interacted with desmoglein 1, desmoplakin, and keratins in the yeast two-hybrid system. Overexpression of the armadillo repeat domain induced a striking dominant negative phenotype with the formation of filopodia and long cellular protrusions, where plakophilin 1 colocalized with actin filaments. This phenotype was strictly dependent on a conserved motif in the center of the armadillo repeat domain. Our results demonstrate that plakophilin 1 contains two functionally distinct domains: the head domain, which could play a role in organizing the desmosomal plaque in suprabasal cells, and the armadillo repeat domain, which might be involved in regulating the dynamics of the actin cytoskeleton.

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Available from: Christof Haffner, Mar 24, 2015
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    • "In HeLa cells, the protein predominantly localized to the cell surface and, therefore, interaction with GTF2IRD1 was not tested by co-IP. However, the family member PKP1 is known to localize more readily to the nucleus (Hatzfeld et al. 2000) and we considered the possibility that the interaction of GTF2IRD1 with PKP family members may be conserved. To address this question, the prey vector pGADT7 containing the PKP1 open reading frame was co-transformed with GTF2IRD1 and an interaction in yeast was verified (Supplementary Material, Fig. S2). "
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    ABSTRACT: GTF2IRD1 is one of the three members of the GTF2I gene family, clustered on chromosome 7 within a 1.8 Mb region that is prone to duplications and deletions in humans. Hemizygous deletions cause Williams-Beuren syndrome (WBS) and duplications cause WBS duplication syndrome. These copy number variations disturb a variety of developmental systems and neurological functions. Human mapping data and analyses of knockout mice show that GTF2IRD1 and GTF2I underpin the craniofacial abnormalities, mental retardation, visuospatial deficits and hypersociability of WBS. However, the cellular role of the GTF2IRD1 protein is poorly understood due to its very low abundance and a paucity of reagents. Here, for the first time, we show that endogenous GTF2IRD1 has a punctate pattern in the nuclei of cultured human cell lines and neurons. To probe the functional relationships of GTF2IRD1 in an unbiased manner, yeast two-hybrid libraries were screened, isolating 38 novel interaction partners, which were validated in mammalian cell lines. These relationships illustrate GTF2IRD1 function, as the isolated partners are mostly involved in chromatin modification and transcriptional regulation, whilst others indicate an unexpected role in connection with the primary cilium. Mapping of the sites of protein interaction also indicates key features regarding the evolution of the GTF2IRD1 protein. These data provide a visual and molecular basis for GTF2IRD1 nuclear function that will lead to an understanding of its role in brain, behaviour and human disease.
    Human Genetics 08/2015; 134(10). DOI:10.1007/s00439-015-1591-0 · 4.82 Impact Factor
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    • "PKPs are located at the membrane, in the cytoplasm, and in the nucleus. PKPs act as a structural scaffold for desmosome formation, clustering, and maturation through interactions with the desmosomal cadherins, desmoplakin, intermediate filaments , and perhaps actin (Hatzfeld et al., 2000; Chen et al., 2002; Bonne et al., 2003). In addition, their association with nuclear and regulatory proteins suggests that they can function as signaling modulators important for tissue differentiation, cell–cell contact formation, and migration (Green and Simpson, 2007; Bass-Zubek et al., 2009; Getsios et al., 2009; Godsel et al., 2010; Green et al., 2010; Thomason et al., 2010). "
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    ABSTRACT: Plakophilin 2 (PKP2), a desmosome component, modulates the activity and localization of the small GTPase RhoA at sites of cell-cell contact. PKP2 regulates cortical actin rearrangement during junction formation, and its loss is accompanied by an increase in actin stress fibers. We hypothesized that PKP2 may regulate focal adhesion dynamics and cell migration. Here we show that PKP2-deficient cells bind efficiently to the extracellular matrix, but upon spreading display total cell areas ∼30% smaller than control cells. Focal adhesions in PKP2-deficient cells are ∼2 × larger and more stable than in control cells, and vinculin displays an increased time for fluorescence recovery after photobleaching. Furthermore, β4 and β1 integrin protein and mRNA expression is elevated in PKP2-silenced cells. Normal focal adhesion phenotypes can be restored in PKP2-null cells by dampening the RhoA pathway or silencing β1 integrin. However, integrin expression levels are not restored by RhoA signaling inhibition. These data uncover a potential role for PKP2 upstream of β1 integrin and RhoA in integrating cell-cell and cell-substrate contact signaling in basal keratinocytes necessary for the morphogenesis, homeostasis, and reepithelialization of the stratified epidermis.Journal of Investigative Dermatology advance online publication, 25 July 2013; doi:10.1038/jid.2013.266.
    Journal of Investigative Dermatology 06/2013; 134(1). DOI:10.1038/jid.2013.266 · 7.22 Impact Factor
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    • "Recent studies have shown a functional link between the desmosomal components and the actin cytoskeleton. For instance, plakophi- lin-1, a p120 related desmosomal plaque protein, has been demonstrated to associate with actin and is capable of inducing filopodium formation [18]. Similarly, plakophilin-2 is also reported to be involved in the regulation of Rho activity and actin organisation during intercellular junction assembly [19]. "
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    ABSTRACT: Desmoglein 3 (Dsg3), a member of the desmoglein sub-family, serves as an adhesion molecule in desmosomes. Our previous study showed that overexpression of human Dsg3 in several epithelial lines induces formation of membrane protrusions, a phenotype suggestive of Rho GTPase activation. Here we examined the interaction between Dsg3 and actin in detail and showed that endogenous Dsg3 colocalises and interacts with actin, particularly the junctional actin in a Rac1-dependent manner. Ablation of Rac1 activity by dominant negative Rac1 mutant (N17Rac1) or the Rac1 specific inhibitor (NSC23766) directly disrupts the interaction between Dsg3 and actin. Assembly of the junctional actin at the cell borders is accompanied with enhanced levels of Dsg3, while inhibition of Dsg3 by RNAi results in profound changes in the organisation of actin cytoskeleton. In accordance, overexpression of Dsg3 results in a remarkable increase of Rac1 and Cdc42 activities and to a lesser extent, RhoA. The enhancements in Rho GTPases are accompanied by the pronounced actin-based membrane structures such as lamellipodia and filopodia, enhanced rate of actin turnover and cell polarisation. Together, our results reveal an important novel function for Dsg3 in promoting actin dynamics through regulating Rac1 and Cdc42 activation in epithelial cells.
    Experimental Cell Research 07/2012; 318(18):2269-83. DOI:10.1016/j.yexcr.2012.07.002 · 3.25 Impact Factor
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