Plakophilins 2 and 3 are involved in desmosome formation in the intestine

Abstract

The integrity of desmosomes is crucial to maintain intestinal epithelial barrier (IEB) function in health and disease. In addition to adherens junctions, desmosome‐dependent intercellular adhesion between cells is provided by the cadherins, Desmoglein2 (DSG2) and Desmocollin2 (DSC2) in the intestine. Both are tethered to the intermediate filament system via different adaptor proteins, such as the Plakophilins (PKPs). The specific role of the different isoforms of PKP 1–3 has been recognized in different organs. However, there is a wide lack of literature regarding the existence of the PKPs and their critical role in the intestine. Accordingly, it remains unclear whether PKPs play a specific role for IEB stabilization. Therefore, the aim of this study was to perform a basal characterization of intestinal PKPs.To analyze the expression pattern of the different PKP isoforms, immunostaining, qRT‐PCR and Western blotting were used. In healthy human mucosa that derived from surgical specimens, PKP2 and PKP3, but not PKP1, were present at the cell borders of tissue sections. In co‐staining experiments with DSG2, both PKP2 and PKP3 were found to be present at desmosomal junctions. The expression of PKP2 and PKP3 but not PKP1 was confirmed on the RNA level by qRT‐PCR and on the protein level by Western Blot analysis. Similar results were obtained in enteroids generated from human mucosa and in Caco2 cells which are commonly used as models to investigate the mechanisms underlying changes of IEB showed a similar expression pattern of PKPs. Interestingly, in Caco2 cells that are deficient for DSG2, a compensatory upregulation of PKP2 RNA and total protein levels was detected when compared to Caco2 WT cells whereas no changes were evident for PKP3 in DSG2‐deficient cells. Stimulation of Caco2 WT cells with Tumor necrosis factor α (TNFα) resulted in a loss of IEB as revealed by measurements of transepithelial resistance. This was associated with a loss of DSG2, PKP2 and PKP3 at the cell borders. However, while total protein levels of PKP2 and DSG2 were unaltered under these conditions, PKP3 protein levels were upregulated.In summary, for the first time, we provide evidence that PKP2 and 3 but not PKP1 are expressed in human intestinal epithelial cells as well as in human enteroids and Caco2 cells. Compensatory upregulation of PKP2 following loss of DSG2 suggests a functional role for PKP2 to stabilize desmosomal integrity. Furthermore, since TNFα treatment was accompanied with an upregulation of PKP3, a potential role of PKP3 during inflammatory processes can be hypothesized.