Polarization is a universal trait of eukaryotic cells and every cell acquires features of polarization – either transiently or stably – during its life. In some cases cell polarity is reflected by an asymmetric distribution of molecules within the cells and as such is not visible unless these molecules are visualized. For example, in the fertilized egg of C.elegans certain proteins localize exclusively to the anterior pole (shown in red in Fig.1A) and others exclusively to the posterior pole (shown in green). Epithelial cells, as another example, have distinct membrane domains, an apical (shown in green in Fig. 1D) and a basolateral membrane (shown in blue), which differ in the composition of proteins; this is also referred to as membrane polarity. In other cases, polarization reflects morphological asymmetry and is illustrated for example by the morphological changes in cytotoxic T lymphocytes (CTL) during lysis of target cells (Fig.1B) or by the presence of a single axon and several dendrites in neurons (Fig.1C).
Clearly, polarization is required for the cell to function properly. For example, the presence of several dendrites and a single axon in neurons determines the unidirectional flow of signals. Also, the formation of different membrane domains in epithelial cells with distinct identities is required for the directional uptake of nutrients from the lumen of the gut by intestinal epithelial cells. The establishment of cell polarity is thus not only required for the functioning of the individual cell but is a prerequisite for the development of whole organs and consequently the entire organism.
Cell Polarity Proteins
In the last few years the molecular mechanisms underlying cell polarity formation have attracted much interest in cell biology. Several highly conserved proteins have turned out as master regulators of cell polarity. One group of cell polarity proteins consists of PAR proteins (PAR-1 to PAR-6) with a complex consisting of PAR-3, aPKC and PAR-6 forming a functional unit. The PAR proteins have originally been identified in C.elegans in a genetic screen for genes regulating the partitioning of cytoplasmic granules (par stands for partitioning-defective), and PAR-3 and PAR-6 together with atypical PKC form a functional unit (the PAR-3 – aPKC – PAR-6 complex). Meanwhile the PAR proteins have turned out to be used by a large variety of cell types as diverse as fertilized eggs, epithelial cells, neurons, astrocytes or leukocytes, and to be involved in processes like membrane specification, tight junction formation, axon specification or cell migration. In part, the ability to regulate such diverse functions resides in the ability of the PAR-3 – aPKC – PAR-6 complex to influence microtubule polymerization and stability.
Epithelial and endothelial cells
We have focussed our research interest on the mechanisms underlying the formation of cell polarity in epithelial and endothelial cells of vertebrates. Epithelial cells often form monolayered sheets of cells that line surfaces of organs and form the interface between the interior and the exterior of the organism, for example in the respiratory tract, the gastro-intestinal tract, or the genito-urinary tract. Endothelial cells are specialized epithelial cells that line the inner surface of the blood vessels. In both cell types, the plasma membrane is divided in two domains: the region of the membrane which is "free", i.e. not in contact with another cell, is defined as the apical domain, the membrane regions which are in contact with adjacent cell or with the underlying extracellular matrix are defined as the basolateral domain. These domains are separated by cellular junctions, in particular the tight junctions in vertebrates, which maintain this membrane polarity by preventing the intermixing of the two domains. Understanding the mechanisms regulating the formation of tight junctions is therefore also critical to the understanding of membrane polarity in epithelial and endothelial cells.
A number of integral membrane proteins has been identified at the tight junction. Many of these mediate adhesion between neighbouring cells through homophilic interactions and help to anchor the individual cell within the multicellular sheet. They all associate with cytoplasmic proteins and are frequently linked through these interactions with the actin cytoskeleton. Although the biological role of these interactions is still unknown in many cases, it became clear that they are far from being static but rather highly dynamic and subject to multiple modes of regulation. One principle that has emerged during the last few years is that the integral membrane proteins through their association with cytoplasmic proteins help to organize large, multimolecular signaling complexes at the tight junction.
Cell adhesion molecules in cell polarity
We are primarily interested in two families of cell adhesion molecules, Junctional Adhesion Molecules (JAMs) and Cadherins, and their role in cell-cell contact and cell polarity formation.
Junctional Adhesion Molecules (JAMs)
JAMs comprise a small subfamily within the immunoglobulin superfamily consisting of three members (JAM-A, -B, -C). They all undergo homophilic interactions and are expressed by different cell types including epithelial cells, endothelial cells, leukocytes and in cells of the reproductive system (Sertoli cells, spermatids). In epithelial and endothelial cells, they are localized at tight junctions. We have focussed our work on the identification of intracellular binding partners of JAMs and have identified several proteins including ZO-1, AF-6/afadin and PAR-3. These proteins have in common one or multiple protein-protein-interaction domains like PDZ domains or guanylate-kinase domains, and they serve to organize the formation of multiprotein complexes. Thus, one major function of JAMs probably resides in the recruitment of specific protein complexes including cell polarity protein complexes to sites of cell-cell adhesion.
VE-cadherin belongs to cadherin superfamily, mediates homphilic binding and is expressed specifically in endothelial cells where it localizes to adherens junctions. It mediates cell-cell adhesion and as in the case of other classical cadherins its cytoplasmic tail binds β-catenin and p120ctn. We have recently identified a cell polarity protein complex in endothelial cells which is associated with VE-cadherin through direct interactions of both PAR-3 and PAR-6 with VE-cadherin. As opposed to the hitherto known PAR protein complex described in epithelial cells, the VE-cadherin-associated PAR complex lacks aPKC. The biological role of this PAR complex in endothelial cells is a major focus of our present work.
Mandicourt, G., Iden, S., Ebnet, K., Aurrand-Lions, M. and Imhof, B.A. (2007). JAM-C regulates tight junctions and integrin-mediated cell adhesion and migration.J. Biol. Chem. 282: 1830-1837.
|Iden, S., Rehder, D., August, B., Suzuki, A., Wolburg-Buchholz, K., Wolburg, H., Ohno, S., Behrens J., Vestweber, D. and Ebnet, K. (2006). A distinct PAR complex associates physically with VE-cadherin in vertebrate endothelial cells. EMBO Rep. 7: 1239-1246.|
|Rehder, D., Iden, S., Nasdala, I., Wegener, J., Meyer zu Brickwedde, M.K., Vestweber, D. and Ebnet, K. (2006). Junctional adhesion molecule-A participates in the formation of apico-basal polarity through different domains. Exp. Cell Res. 312: 3389-3403.|
|Gliki, G., Ebnet, K., Aurrand-Lions, M., Imhof, B.A. and Adams, R.H. (2004). Spermatid differentiation requires the assembly of a cell polarity complex downstream of junctional adhesion molecule C. Nature 431: 320-324.|
|Ebnet, K., Suzuki, A., Ohno, S. and Vestweber, D. (2004). Junctional adhesion molecules (JAMs): More molecules with dual functions? J. Cell Sci. 117: 19-29.|
|Ebnet, K., Aurrand-Lions, M., Kuhn, A., Kiefer, F, Butz, S., Zander, K., Meyer zu Brickwedde, M.K., Suzuki, A., Imhof, B.A. and Vestweber, D. (2003). The junctional adhesion molecule (JAM) family members JAM-2 and JAM-3 associate with the cell polarity protein PAR-3; A possible role for JAMs in endothelial cell polarity. J. Cell Sci. 116: 3879-3891.|
|Ebnet, K., Suzuki, A, Horikoshi, Y, Tomonori, H., Meyer zu Brickwedde, M.K., Ohno, S. and Vestweber, D. (2001). The cell polarity protein ASIP/PAR-3 directly associates with junctional adhesion molecule (JAM). EMBO J. 20: 3738-3748.|
|Ebnet, K., Schulz, C.U., Meyer zu Brickwedde, M.K., Pendl, G.G. and Vestweber, D. (2000). Junctional adhesion molecule interacts with the PDZ domain-containing proteins AF-6 and ZO-1. J. Biol. Chem. 275: 27979-27988.|