The area composita of adhering junctions connecting heart muscle cells of vertebrates.: V.: The importance of plakophilin-2 demonstrated by small interference RNA-mediated knockdown in cultured rat cardiomyocytes

In the adult mammalian heart, the cardiornyocytes are connected by large polar arrays of closely spaced or even fused composite, plaque-bearing adhering junctions,(areae contpositae, ACs), in a region usually termed intercalated disk, (ID). We have recently reported that during late embryogenesis and postnatally these polar assemblies of ACjunction structures are gradually formed as replacements of distinct embryonal junctions representing desmosomes and Jasciae adhaerentes which then may amalgamate to the fused AC structures, in some regions occupying more than 90% of the total ID area. Previous gene knockout results as well as mutation analyses of specific human cardi'omyopathies have suggested that among the various AC constituents, the desmosomal plaque protein, plakophilin-2, plays a particularly important role in the formation, architectural organization and stability of these junctions interconnecting mature cardiomyocytes. To examine this hypothesis, we have decided to study losses of - or molecular alterations in such AC proteins with respect to their effects on myocardiac organization and functions. Here we report that plakophilin-2 is indeed of obvious importance for myocardial architecture and cell-cell coupling of rat cardiornyocytes growing in culture. We show that siRNA-mediated reduction of the cardiomyocyte content of plakophilin-2 but not of some other major plaque components such as desmoplakin results in progressive disintegration - and losses - of AC junction structures and that numerous variously sized vesicles appear, which are plaque protein-associated as demonstrable by immunofluorescence and immunoelectron microscopy. The importance of plakophilin-2 as a kind of organizer protein in the formation, stabilization and functions of the AC structure and the ID architecture is discussed in relation to other junction proteins and to causes of certain cardiornyopathies. (c) 2007 Elsevier GmbH. All rights reserved.