Genes And Proteins Involved In Celltocell And Cellmatrix Adhesion

Epithelial cells adhere to each other and interact with each other through several types of contacts (Figure 9.2). Morphologically distinct and functionally important contacts include adherens junctions, gap junctions, and tight junctions (occluding junctions).

Occluding junctions seal epithelia and define the apical and lateral membrane compartments of an epithelial cell. Therefore, their loss in a carcinoma cell is associated with the loss of cell polarity.

Claudin Ocoludin

Claudin Ocoludin

Figure 9.2 Cell-cell- and cell-matrix-contacts of epithelial cells A: Tight junctions; B: Adherens junctions; C: Hemidesmosomes; D: Integrin / extracellular matrix contacts. Different integrins prefer different matrix proteins.

Adherens junctions are arranged in a belt-like configuration (hence 'belt desmosomes') between adjacent epithelial and are intracellularly connected to actin filaments. The proteins actually mediating homotypic interactions between adjacent epithelial cells are cadherins, of which E-Cadherin is a typical representative (Figure 9.3). Interaction between E-Cadherin molecules is Ca2+-dependent. On the cytoplasmic surface of the cell membrane, E-Cadherin is linked to actin filaments by a-Catenin and P-Catenin. Down-regulation or mutation of E-Cadherin is frequent in human cancers and often occurs during tumor progression. In some invasive tumors, E-Cadherin is replaced by other members of the family with adhesion properties more suitable for a migrating cell, e.g. N-Cadherin. This is labelled a 'cadherin switch'.

Spot desmosomes are connected to cytokeratin filaments in the epithelial cell by desmoplakin and plakoglobins. The actual contact between cells is again made by specialized cadherins.

Gap junctions also contribute to adhesion between cells, but primarily are communication channels that connect cells of the same type in epithelia or in other tissues, e.g. the nervous system. They allow the passage of small molecular weight molecules (« 1000 Da), in particular of cations such as Ca2+ and K+, but also of other small signaling molecules such as cyclic AMP. Gap junctions are formed from one connexon in each partner cell, which consists of six connexin molecules. Different connexins are expressed in a tissue-specific fashion accounting for the specificity of the interaction. Gap junctions are temporarily closed, when cells separate from their neighbors for division. Closure is regulated by phosphorylation of connexins. Gap junctions are also sensitive to high Ca2+ levels and accordingly shut when cells become apoptotic or necrotic. In cancers, gap junctions become as a rule inactive during tumor progression and tissue-specific connexin expression is down-regulated.

In addition to adhering to each other, at least the basal cells in an epithelium are attached to the basement membrane. This contact is made through integrins (^ Figure 6.8) which bind fibronectin and laminin in the extracellular matrix. On the cytoplasmatic side they are attached to actin filaments via a-actinin, vinculin and talin. Integrins are heterodimers consisting of each one a and P subunit. There are >17 a- and >8 P subunits in humans. A typical integrin of an epithelial basal cell is a2p1. A specific integrin, a6p4, is a constituent of hemidesmosomes. These also attach epithelial cells to the basement membrane matrix, but are attached to the cytokeratin cytoskeleton on the cytoplasmatic face. The composition of integrins is cell-type specific, and changes in migratory cells. For instance, cells of the hematopoetic lineage down-regulate their expression of a4p1 integrin, when they have become differentiated and leave the bone marrow. Tumor cells often express different integrins than their normal counterparts. Down-regulation of integrins accounts, e.g., for the failure of chronic myelogenous leukemia cells to remain in the bone marrow sufficiently long for complete differentiation (^10.4). In carcinomas, too, changes in integrin composition may be essential for invasion and metastasis, with some integrins favorable and others inhibitory for invasion.

Figure 9.3 Structure and interactions of E-Cadherin See text for further explanation

Cell-to-cell adhesion and cell-matrix adhesion elicit signals within the cell, e.g. integrins through protein kinases like FAK, ILK, and SRC family tyrosine kinases that influence several pathways regulating cell growth and survival (^6.5). In epithelial cells, lack of attachment tends to cause anoikis, a specific kind of apoptosis. Alterations in signaling as a consequence of altered adhesion in carcinoma cells must therefore be compensated. This may be the reason, why altered SRC expression is a common finding in cancers.

Importantly, the ability to migrate is not simply acquired by loss of adhesive properties. Instead, cell migration requires a dynamic pattern of adhesion contacts that may overall be more complex than in a cell residing in a tissue. A single cell migrating through a tissue needs to adhere to the extracellular matrix to just the right extent, i.e. sufficiently tightly to pull itself through the tissue, but not so tightly as to become unable to extricate itself. Moreover, if a net movement is to be achieved, attachments need to be established at a leading edge and be broken at a trailing edge by well-localized proteolysis, and cytoskeletal contractions must be coordinated. Localized activation of proteases at certain points on the cell surface is therefore a prerequisite for migration in a tissue (^9.3). Some normal and tumor cells also move by a more amoeboid mode with less attachment and less activity of the actin cytoskeleton.

In fact, carcinoma cells more often migrate as cell clusters or cell files, or even expand like cell sheets during fetal development. These modes of migration nevertheless require reorganization of cell-to-cell adhesion and the cytoskeleton compared to cells in a resting epithelium. For instance, they may be accompanied by changes in the integrin isotypes and in the type of cadherin present. Some carcinomas also undergo a morphological change to a more mesenchymal phenotype, called an 'epithelial-mesenchymal transition' (sometimes abbreviated as EMT). This allows them to invade and migrate as single cells, which is otherwise more typical of hematological and soft tissue cancers. However, cell-to-cell and cellmatrix adhesion remain dynamic in tumor tissues, as a rule, and the EMT can be reversible.

The course of E-Cadherin expression in the course of prostate cancer development illustrates the dynamic nature of changes in cell adhesion in cancers. E-Cadherin becomes down-regulated during invasion of prostate cancer cells into the stromal tissue compartiment, but is re-expressed in many metastases. A modulation of gene expression in this fashion is obviously more efficiently achieved by epigenetic than by genetic alterations. So, the promotor of the CDH1 gene encoding E-Cadherin is hypermethylated in some carcinomas in a highly variable fashion that is associated with a variable expression of the protein.

In contrast, cancers with a diffuse-type growth pattern down-regulate specific cell adhesion molecules irreversibly. Again, E-Cadherin can serve as an example. Diffuse-type gastric cancer is characterized by highly invasive small groups of loosely adherent undifferentiated tumor cells. In this cancer type, E-Cadherin is regularly inactivated by mutation and deletion of the gene. In fact, germ-line mutations in CDH1 are responsible for rare familial cases of this cancer (^17.2).

Mutations in E-Cadherin are among the few genetic changes that specifically alter cell adhesion molecules in carcinoma cells. A much wider range of changes on the surface of tumor cells (Table 9.1) are achieved by epigenetic mechanisms. These include aberrant methylation of genes like CDH1 and others associated with more or less strong down-regulation of expression. In other cases, up-regulation, altered splicing, or altered processing of cytoskeletal and membrane proteins are observed. Often, these changes in invasive tumor cells give the impression of a coordinated switch in gene and protein expression towards an 'invasion program' rather than that of an accumulation of alterations in individual genes selected to yield an invasive phenotype.

Nevertheless, some among the myriad changes at the cell surface of cancer cells may be more important than others and may be decisive for invasion and metastasis. In particular, down-regulation of specific genes appears to be required for metastasis in certain cancers, although these genes do not influence the growth of primary tumors. Such genes have been designated 'metastasis suppressor genes'. Most of

Table 9.1. Some cell surface proteins altered in human cancers

Protein(s)

Function

Alteration(s) in cancer

E-Cadherin

homotypic cell adhesion

mutation, down-

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