Cells are independent units of life. However, when a cell becomes part of an organism, it becomes part of a tissue and organ system. Cells in a tissue are joined to each other and to the extracellular matrix (ECM) by cell junctions. There are three kinds of cell junctions: Occluding junctions, anchoring junctions and communication junctions. Occluding junctions occur in the epithelium where the adjacent cells are so tightly sealed together that even small molecules cannot pass. Communicating junctions allow the exchange of chemical and electrical signals between cells. Anchoring junctions are the ones which attach a cell either to its neighbor or to the extracellular matrix. Formation of an anchoring junction requires the cells to stick to each other. Various molecules mediate adhesion between cells following which the cytoskeleton forms a structure around them. The anchoring junctions thus formed can be of four types- desmosomes, hemidesmosomes, focal adhesions and adherens junctions. In order for a proper tissue to form, it is important for the cells of the tissue to bind together. It is equally important to prevent the invasion and binding of other cells. Thus, cell adhesion has to be specific. This is where the cell adhesion molecules come into play (Alberts et al. 2002).
Cell Adhesion Molecules (CAMs)
Adhesion of cells to other cells and to the ECM is mediated by certain proteins on the surface of the cells called cell adhesion molecules or CAMs. CAMs can be divided into two kinds: Ca2+ -dependent and Ca2+ -independent. There are four major CAMs: cadherins, integrins, selectins and the immunoglobulin (Ig) superfamily
Cadherins are a family of proteins that are responsible for Ca2+ dependent cell adhesion. The cadherin superfamily consists of five different members. The type-I cadherins and type-II cadherins are also called the classical cadherins. The type-I cadherins are classified based on the tissue in which they were first discovered. The cadherins present on epithelial cells are called E-cadherins and those on neurons and muscle cells are called N-cadherins. Cadherins present on placental and epidermal cells are called P-cadherins. The type-II cadherins have overlapping expression patterns. Apart from these there are a variety of non-classical cadherins present in human cells. The third type of cadherins is the desmosomal cadherins that are the adhesion molecules responsible for the formation of the intercellular desmosomal junctions. There are two types of desmosomal cadherins: desmocollein and desmoglein (Garrod et al. 2002). Protocadherins belong to a subfamily of the cadherin super-family that exhibit weak adhesive properties Frank and Kemler 2002). Apart from these there are other varieties of cadherins. One example is the T-cadherin that is present in the nerve and muscle cells; it lacks a transmembrane domain but remain attached to the plasma membrane by a glucosylphosphatidylinositol anchor (Alberts et al. 2002).
Cadherin proteins have an extracellular domain and a transmembrane domain by which they are anchored to the plasma membrane. The extracellular domain is composed of five or six cadherin repeats. The cadherin repeats are structurally similar to Immunoglobulin domains. Calcium ions are present in between every cadherin repeat. The Ca2+ ions make the extracellular domain rigid and help in its binding to the cadherins of a neighboring cell (see fig 1).
E-cadherin is the most extensively characterized member of the cadherin superfamily. E-cadherin is composed of a transmambrane domain, an ectodoamin and a cytoplasmic domain. The ectodomain contains five cadherin repeats of which the last one has four cysteins. The cytoplasmic domain of E-cadherins contains two sub-domains: the juxtamembrane domain and the beta-catenin binding domain. E-cadherins play a crucial role during development. During the eight-celled stage of embryo development, the loosely attached blastomere cells become tightly adhered to each other with the help of E-cadherins. The specificity of E-cadherins for this stage was demonstrated by the mutant expression of N-cadhrins in cells that lack E-cadherins. Such cells formed neuroepithelium and cartilage instead of epithelium. E-cadherins are also important during development in a process known as cadherin switching. During the formation of the neural tube, the cells in the neural tube lose E-cadhrins and gain other cadhrins such as N-cadherins. The overlying ectodermal cells express E-cadherins until the nueral crest cells migrate away. The switching of the cadherins is important in the development; if N-cadherins are over-expressed in the neural crest cells, the cells are unable to migrate away from the neural tube (Roy and Berx, 2008).
Types of adhesions
Adhesion mediated by cadherins can be of two types: homophilic adhesion and heterophilic adhesion. The homophilic adhesion mechanism was demonstrated in L-cells, a line of cells that do not express cadherins. When L-cells were transfected with different cadherin genes and mixed together, cells expressing the same type of cadherins bound together suggesting the homophilic binding preference of cadherin molecules. Apart from this homophilic adhesion mechanism, cadherins bind to some other specific adhesion molecules such as integrin E7 and a bacterial protein known as internalin (Roy and Berx, 2008).
Catenins-Cadherin associated molecules
Cadherins join cells together at cell junctions and thus indirectly connects the actin cytoskeleton molecules of the cells involved. The cytoplasmic domain of the cadherin molecules are joined to actin filaments by a group of proteins known as catenins (see Fig 2). Catenins were discovered by immunoprecipitation of cadherin molecules using anti-cadhein antibodies. There are three major types of catenins: -catenins, -catenins and plakoglobins (-catenins). The cytoplasmic domains of the classical cadherin molecules bind to -cateinins and plakoglobins. These catenins contain a conserved Armadillo domain consisting of nine to twelve repeats that fold into a conserved structure enabling them to interact with cadherin molecules. The -catenins lack an Armadillo domain and thus fail to bind to cadherins. The however act as a connecting link between the other catenin molecules and the actin cytoskeleton (Shapiro and Weis 2009).
Integrins are transmembrane cell-adhesion molecules that link the extracellular matrix to the cytoskeleton. Integrins act as receptor molecules for the binding of extracellular matrix proteins. Integrins bind to their ligands with a weak adhesion, but are present in a huge number. Thus the interaction mediated by integrins is dependent on a large quantity of weak interactions. This kind of interaction is ideal since it prevents the cell from being immovably attached to the extracellular matrix. Apart from just anchoring the cell to the extracellular matrix, integrins act as receptors carrying signals from the extracellular matrix to the inside of the cell (Alberts, 2002).
Integrins exist as heterodimers and are composed of an and a subunit. Both the subunits are composed of a membrane-spanning domain and a cytoplasmic tail. The extracellular domain of the subunit has five parts: a -propeller, a thigh and two calf domains. The -propeller domain contains calcium binding sites that are crucial for ligand binding. The extracellular domain of the -subunit has seven parts. The -1 domain is followed by a hybrid domain and a plexin-semaphorin-integrin domain. These are followed by four epidermal growth factor domains and a -tail. The transmembrane domains of the two subunits associate to form an inactive receptor. The cytoplasmic domains of both the subunits are thought to be flexible and forming transiently interacting structures. Most integrins bind to actin filaments through their cytoplasmic tail domains with the exception of 64 integrins, which bind to intermediate filaments (Campbell and Humphries 2011).
Integrins bind to extracellular ligands in the matrix through their extracellular domain. Based on the ligand to which they bind, integrins are classified into laminin-binding integrins, collegen-binding integrins, leukocyte-binding integrins and RGD-binding integrins. The binding of the ligands is mediated by various ions like Ca2+, Mn2+ and Mg2+. After the integrins bind to extracellular ligands through their ectodomains, the cytoplasmic domains interact with various intracellular proteins like talin, actinin and filamin. These proteins serve as anchors connecting the integrin proteins to the actin cytoskeleton. The deletion of the cytoplasmic domain results in integrins that can bind to their ligands but fail to form proper adhesions (Campbell and Humphries 2011). This phenomenon emphasizes the importance of the binding of integrins to cytoskeletons for proper cell-matrix interaction (see Fig 3).
Inside –out signalling for intergin regulation
Singals generated inside the cells regulate the activity of intergins. G-protein coupled receptors (GPCRs) play a role in signalling cascade involved in the regulation of integrin molecules. The activation of integrins is mediated by phoshorylation triggered by G-protein coupled receptor signalling. Similarly the disruption of the interaction of and subunits in their cytoplasmic domains is mediated by chemokines via GPCRs. Apart from GPCRs the cytoplasmic protein talin also mediates the activation of integrins through the disruption of - interaction. Binding of talin disrupts the interaction and causes a conformational change in the ectodomains. This induces the activity of integrins. Two models have been proposed to explain this change in activity. The first is called the “deadbolt” model. According to this model, an activated integrin maintains a bent conformation that is characteristic of its inactive stage. However, movements in the transmembrane domains mediate a disruption in the interaction between the two subunits thereby activating the receptor. The second model is called a “switchblade model” in which the transmemebrane domains of the two subunits dissociate and mediate the separation of an EGF repeat in the -subunit. This separation causes the head region to stretch outward (Takada et al. 2007)
Signaling mediated by integrins
Integrins mediate a variety of signaling cascades. Binding of ligands to the extracellular domains induce conformation changes in the integrin molecules. These conformation changes send signals to the inside of the cell and mediate various cellular processes. Signaling by integrins activate MAPK pathways in most cases. There are two major mechanisms by which integrins activate the MAPK pathway: the direct mechanism and the indirect mechanism. The direct signaling cascades activated by integrins are mediated by an intracellular tyrosine-kinase receptor molecule known as the Focal Adhesion Kinase (FAK). The intracellular protein talin recruit the FAKs to the point of cell-matrix contact. Cross-phosphorylation of FAKs induce the binding of another family of tyrosine kinases called Src kinases. Src kinases further phosphorylate and activate FAKs. FAKs then phoshorylate other intracellular molecules and thus convey the signal to the inside of the cell. Other than the FAK-dependent signaling, integrin interacts with the kinase Fyn and an adaptor protein called Shc. This interaction is brought about by the protein caveolin-1. Binding of integrin induces Fyn to phosphorylate Shc. This phosphorylation then activates the signaling cascade. During indirect signaling pathways, integrins engange in cross-talk with other cellular signaling cascades.
Intergins interact with other signaling cascades inside the cell and regulate the activation and expression of each other. Integrin signaling and Ras/MAPK signaling together regulate the process of cell proliferation. The importance of integrin signaling for the survival of the cell is demonstrated by the fact that cell lines that lose contact with the extracellular matrix fail to proliferate normally and sometime even undergo programmed cell death (Takada et al. 2007).
Selectins are adhesion molecules that mediate cell-cell adhesions in the bloodstream. Selectins are cell-surface proteins that cause temporary Ca2+-dependent adhesions between the cells in the bloodstream. There are three types of selectins: L-selectins, P-selectins and E-selectins. L-selectins are present on white blood cells and P-selectins are present on platelet cells. E-selectins are present on endothelial cells. Selectins are transmembrane molecules contain an extracellular domain, an epidermal growth factor motif and several consensus repeat sequences (Tedder, 1995).
Selectins bind to a variety of ligands of which the P-selectin glycoprotein ligand 1(PSGL-1) is the most widely characterized. Recruitment of leukocytes during inflammation involves the capture, rolling migration and adhesion of the blood cells. Selectins play a crucial role in the signaling involved in leukocyte recruitment. In monocytes, binding of PSGL-1 to selectins enable them to leave the vascular system. In mouse models, the reduction of P-selectins reduces the adhesion of monocytes to the epithelium. P-selectins on activated platelets bind to PSGL-1 on leukocytes and monocytes. This binding helps the migration of leukocytes. Selectins induce a weak adhesion of the white blood cells to the endothelium. This adhesion enables the blood cells to roll along the endothelial surface until the activation of the integrins. Acvtivated integrins help the blood cells to strongly adhere to the endothelium. Selectins and integrins thus together help the adhesion of blood cells to the endothelium (Claus 2003).
Members belonging to the immunoglobulin superfamily mediate calcium-independent cell-cell adhesion. These proteins are characterized by Ig-like domains. The best-characterized Ig adhesion molecule is the N-CAM (neural cell adhesion molecule) that is present in various cells. Human cells encode a single N-CAM gene that undergoes alternative splicing to produce more than twenty different proteins. The cell adhesions mediated by Ig molecules are weaker than those by cadherins and play a role in the extension of these interactions especially during development. During the development of pancreas in mice, the islet of Langerhans forms by a process of cell aggregation and cell-sorting. It has been discovered that though cadherin-mediated binding is important for cell aggregation, cell sorting requires N-CAM. Mutations in the gene encoding N-CAM cause developmental defects in mice emphasizing its importance. In Drosophila, mutations in the N-CAM-like protein FAS3 disable the nerve cells’ ability to recognize muscle targets (Alberts et al. 2002, takada et al 2007).
The extracellular domain of N-CAMs is composed of two components. There are five Ig-like repeats that are connected to each other by disulfide bonds. This part is followed by two fibronectin type-III domains. The Ig-like domains are responsible for homophilic binding to other Igs while the fibronectin domains are responsible for signaling mechanism. Apart from the extracellular domain, Igs have a transmembrane domain that anchors them to the plasma membrane (Weledji and Assob, 2014). Most Ig-like molecules bind to each other through a homophilic mechanism. Some molecules like the intracellular adhesion molecules (ICAMs), however, engage in a heterophilic binding mechanism with integrins (Alberts et al. 2002, Cambpell and Humphries 2011).
Ig-like adhesion proteins are also involved in transmitting extracellular signals to the inside of the cell and mediating cellular processes. In neurons, N-Cam molecules associate with the intracellular tyrosine kinase Src. Src then phosphorylates various other intracellular molecules thereby transmitting signals from the outside of the cell to the interior.
Nectins are a specific class of Ig-like CAMs. They engage in homophilic and heterophilic interaction with each other and other Ig-like molecules. Nectins have a cytoplasmic domain that is connected to the actin cytoskeleton by means of a protein known as afadin. Nectin proteins are composed of an extracellular domain, a transmembrane domain and a cytoplasmic tail.
Nectins and afadins are responsible for the formation of adherens junctions along with cadherin molecules. Nectins form cell adhesions before the recruitment of cadherins. The cell adhesions formed by nectins help in the migration of cadherins to the cell surface and the formation of string junctions. Binding proteins like afadin and -catenin mediate the binding of nectins and cadherins to each other.
Nectins are also responsible for cell-matrix crosstalks through their interaction with integrins. Some forms of nectins interact with integrins at the cell-cell adhesion points. This interaction activates the Src kinase. Src then activates the G-protein Rap1. Rap1 then activates other intracellular molecules and mediate the formation of lamellipodia and filopodia in cells. Nectins play various other roles like cell movement and differentiation, cell polarization and cell survival (Takai et al. 2008).
Role of cell adhesion molecules in cancer
Proper cell-cell adhesion is crucial in maintaining the proper structure and function of tissues. Cancer occurs due to the abnormal proliferation of tissues and the metastasis of the cancerous cells to neighboring tissues. Thus cell-adhesion molecules play a major role in cancer development metastasis, and angiogenesis. All four types of CAMs are involved in these processes. Integrins are expressed in the endothelial cells and play a critical role in cell migration. Cadherins on the vascular endothelium regulates angiogenesis and the integrity of newly-formed blood vessels. Selectin is responsible for the adhesion of leukocytes to activated endothelial cells. It has been shown that in cornea, external addition of selectins can induce the formation of new blood vessels.
Reduced expression of E-cadherins has been associated with breast, pancreas, lung, kidney and prostate cancer. Studies in mouse models suggest that the absence of E-cadherin enables organism to activate the Wnt-induced signaling. Studies have also suggested that E-cadherin is a regulator of pluripotency. E-cahderin interacts with leukemia inhibitory factor and forms teratomas.
Integrins are cell-matrix adhesion molecules that play a role in cell signaling. Binding of integrins to the L1 protein regulates the migration of melanoma cells. During metastasis, integrins and other CAMs help in the adhesion and migration of cancer cells through the blood (Farahani et al 2014).
Cells are joined to each other and to the extracellular matrix by the cell-adhesion molecules. This adhesion is crucial for the proper structure and functioning of tissues. Cell-adhesion is also important in the case of development where the adhesion of a specific tissue to a proper location is vital. In adult systems, cell-adhesion plays a major role in immunity by recruiting and transporting immune molecules to the site of infection or inflammation. In this paper, the role of cell-adhesion molecules in cancer development has also been discussed. Cancer occurs due to abnormal development, division and migration of cells. With the role of cell-adhesion molecules in cancer being studied extensively, the possibility of developing drugs that target CAMs for the treatment of cancer and other diseases is being discussed.
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