Cell Adhesion Molecules(CAM)

Medical Biology

Müjgan Cengiz

Cellular Junctions and Adhesion

• In multi-cellular cells have junctions that occur with:

• 1- Other cells• 2- The Extra-Cellular

Matrix (ECM):

• a network of secreted macromolecular complexes

Figure 19-1 Molecular Biology of the Cell (© Garland Science 2008)

Two main ways in which animal cells are bound together:

•Connective tissue

•Epithelial cells.

Selective Adhesion Determines Specificity of Tissue and Cellular Associations

• Townes & Holtfreter (1950) separated embryonic cells of frogs and combined them together again. To be able to identify the different cell types, they mixed together cells from normal (pigmented) frogs with cells from albino frogs

Identification of CAM through experiment:

Selective Adhesion Determines Specificity of Tissue and Cellular Associations

Antibodies were developed against specific types of CAM.

Antibody treatment of the target cells was found to disturb the cell aggregation.

Identification of CAM through experiment:

• What are the factors that keep the cells together?

• how do the same type of cells recognize each other?

Adhesion

Adhesion = molecules sticking to a substrate

CELL ADESİON MOLECULES

• Cell-cell interactions

• Embryogenesis

• Immunity(migration of immun cells to the inflamation center)

• Cell tissue organ development

• wound healing

• Cancer metastasis

Cell adhesion molecules

• 1-Holding cells together

• 2-Cells detect their extracellular environment through interaction employing a variety of cell adhesion molecules (CAMs)

Cells adhere to each other and to the extracellular matrix through cell-surface proteins called cell

adhesion molecules (CAMs) • Many adhesion molecules are mosaics

of multiple distinct domains• (1) extracellular domain:

– Mediate adhesion

(2) transmembrane domain

• (3) cytosolic domain: • -recruit sets of multifunctional adaptor proteins

• --adaptor proteins: link to cytoskeleton/ signaling molecules

• * outside-in and inside-out effects: connectivity and communication

•

Cell adhesion molecules (CAM)

--Many adhesion molecules are mosaics of multiple distinct domains (1) extracellular domain: mediate adhesion *homotypic adhesion—adhesive interactions between cells of the same

type *homophilic adhesion—a CAM on one cell can directly bind to the same kind of CAM on an adjacent cell.

(2) cytosolic domain: --recruit sets of multifunctional adaptor proteins --adaptor proteins: link to cytoskeleton or signaling molecules

* outside-in and inside-out effects: connectivity and communication

Homophilic Binding- Binding of same kind of molecules in adjacent cells

Heterophilic Binding- Binding to a different kind of molecule on adjacent cells

Linker-dependent Binding-Binding through a secreted linker molecules to other molecules.

1) extracellular domain

The binding of a cell-surface receptor to a secreted ECM molecule immobilized on the substrate.

connects the junction to (micro- or intermediate) filaments

cell adhesion molecules

1- cell adhesion molecules : glicoproteins

2- linker-protein

Cell-cell adhesions can be

• A- tight and long junctions

• e.g. nerve cells in the nerve cells/ the metabolic cells in the liver

• B- relatively weak and transient

• e.g. immune –system cells in the blood

Cell Adhesion Molecules

What are they?

Cell adhesion molecules (CAMS) are cell surface proteins involved in the binding of cells, usually leukocytes, to each other, to endothelial cells, or to the extracellular matrix.

Most CAMS can be placed into one of four general families of proteins:

1) Cadherins

2) Integrins

3) Immunoglobulin (Ig) super family

4) Selectins

JUNCTIONAL ADHESION MECHANISMS Non-JUNCTIONAL CONTACTS

CELL -MATRIXADHESION

CELL -CELLADHESION

basal lamina

actin

NO attachment

plague

CADHERINS

IG-LIKE CAMS

INTEGRINSSELECTINS

adhesion belt(CADHERINS)

desmosomes(CADHERINS)

Gap junctions(connexins)

tightjunctions

focal contacts(integrins)

hemidesm.(integrins) integrins

non-epithelial cellsepithelial cells

• Cell Adhesion also occurs in the blood

• Under most circumstances, all blood cells try to keep from sticking to the wall.

• When a Leukocyte goes on the hunt…– Cell Rolling– Cell Adhesion

Hynes: TiCB: 9:M33 1999

CadherinsCa 2+ -dep. homophilic adhesion functional unit = dimer

Immunoglobin superfamily (CAMs)homophilic or heterophilic

SelectinsheterophilicP selectin + counter-receptor PSGL-1, glycosylated

Integrinsheterodimers, heterophilicbind to ECM, Ig-CAMs, cadherinsadhesion, polarity, migration

Cadh. repeats

Ig and Fn III repeats

lectin repeats

I-CAM

CELL ADHESION MOLECULESCELL-CELL adhesion

Immunobiology, 6th Edition, Janeway, Travers, Walport, and Shlomchik

Some CAMs are Ca2+-dependent,

some others are Ca2+-independent. • Ca2+-dependent

Cadherins, Selectins,

İntegrins

• Ca2+-independent

Ig superfamily

1-CADHERINS

• A family of Ca2+-dependent CAMs

• Ca2+ causes dimerization of Cadherins

• The binding is homophilic-hold cells of one tissue type/subtype together by binding to the same cadherin on a neighbouring cell

• greater the number of cadherins, greater the strength of adhesion

Figure 19-5 Molecular Biology of the Cell (© Garland Science 2008)

Compaction of an early mouse embryo.

At the 8-cell stage they start expressing E-cadherins

1) CA-DEPENDENT CAMs (CELL ADHESION MOLECULES):

a) classic CADHERINS: involved in both junctional and non junctional adhesions

E-, N- and P- cadherins (Epithelial, Nerve, Placenta)

single-pass transmembrane glycoproteins (~700-750 AA s), 5 cadherin repeatsselective adhesion, homophilicdifferential expression during development + morphogenesis

the extracellular side: 5 cadherin repeats of 100 AA, (3 are Ca2+ -binding)

in the absence of Ca >>> rapid proteolysis

the cytoplasmic side:the intracellular attachment proteins: catenins (bind actin) (required for cell-cell adhesion)

Nonclassical cadherines cannot bind actin filaments . They can bind to intermediate filaments and forms desmosomes.

Figure 19-9a Molecular Biology of the Cell (© Garland Science 2008)

Cadherin structure

Extracellular domains of a classical cadherin (C-cadherin)

If Ca2+ is removed, the extracellular part of the protein becomes floppy and is rapidly degraded by

proteolytic enzymes

Ca2+ causes dimerization of Cadherins

Cadherins -Ca-dependent adhesion molecules

Extracellular domains adhere cells together

Link and help assemble the cytoskeleton viaactin or intermediate filaments

Participate in intracellular signalling

Cadherins Are Linked To Actin Cytoskeleton

• The linkage of cadherins to actin filaments.

Different CAMs function in different junctions

Zonula adherens- E-cadherin

•Cadherins interact with actin filaments by CATENINs.

Cadherin-containing junctions connect cells to one another and are linked to

either the actin or IF cytoskeleton

Figure 22-5

, catenins

Localization of sub-types: E-epithelial N-neuronal P-placental VE-endothelial For instance: N-cadherin binds to other N-cadherins, but not to E, P or VE-cadherins This keeps neurons attached to other neurons!

Table 19-3 Molecular Biology of the Cell (© Garland Science 2008)

Embryogenesis & Cadherins

• Expression of specific cadherins accompanies morphogenetic movements during embryogenesis

Embryogenesis & Cadherins

• E-cadherin is the first cadherin expressed during mammalian development. It helps cause compaction, an important morphological change that occurs at the eight-cell stage of mouse embryo development.

• During embryogenesis, production of cadherins promotes cells of similar type to adhere when cells need to migrate to newly growing tissue (e.g. limbs), they lose adhesive properties – cadherins must be endocytosed

•

interior

interior

external

effectors

The changes in cell shape

Cell differentation?Cell- cellAdhesion

GTP

Small GTPaz

Ca2+

GTP

cell

cell

Cadherins and Catenins participate in transduction of extracellular signals and Mediate various cellular response.

2-Integrins:• Mediates Ca+2 dependent

adhesion

Integrin makes cell-substrate interaction

• Ligands: ECM molecules, soluble ligands, CAMs

• ( fibronectin, fibrinogen, ICAM, laminin)

• Two transmembrane glycoprotein subunits: and

• both required for matrix binding

transmembrane linkers of the ECM and the cytoskeleton

bind ligands with low affinity present at very high concentration on the cell surface

Cells both bind to and respond to the ECM via integrins.

3-4 Ca2+ or Mg2+ binding domains on the chain

Diversity of integrins:9 types of , 14 types of ~ 20 different heterodimers identified so far

Integrins are modulated by additional cell-specific factors

Cys

-ric

h

chain

chain

RG

D

RGD” sequence is the specific substrate

Integrins

On the extracellular side integrins bind to the sequence Arg-Gly-Asp found in adhesion molecules including fibronectins

On the intracellular side they bind Vinculin and a-Actinin, these proteins bind to Actin filaments

This dual binding allow cells to move by contracting Actin filaments against the EM

 integrin plays a major role in assembly of the hemidesmosome, or stable  stable anchoring contact.

 

2007 TUS

• Hücrenin, hücreler arası matriks ile etkileşimini sağlayan transmembran yapıdaki adezyon reseptörleri aşağıdakilerden hangisidir? A) Kateninler B) Selektinler C) Kaderinler D) İntegrinler E) Vinkülinler

Aralık 2010

• Aşağıdaki hücre bağlantılarından hangisinin yapısında hücre adezyon molekülü olan integrin yoğun olarak bulunur?A) Zonula occludensB) Zonula adherensC) Macula adherensD) KonneksonE) Hemidesmozom

Integrins Interact with The Cytoskeleton

Integrins are linkers between cytoskeleton and extracellular matrix.

Bind to actin filaments

Need intracellular anchor proteins for bindig to actins.

These intracellular anchor proteins are:

1. Talin

2. α –actinin

3. Filamin

Integrins Are TM Heterodimers ( Combinations) That Mediate Weak Cell-Matrix and Cell-Cell

Interactions

Cell-matrix adhesion is modulated bychanges in the activity and number ofintegrins

De-adhesion factors promote cellmigration and can remodel the cell surface

interactions and the three binding modes of various integrins

1. Integrins bind to ECM proteins via specific amino acid recognition sites

2. Integrins bind to other cell adhesion molecules

3. Integrins promote platelet aggregation through soluble miltivalent mediator molecules

Adherent Cell

Platelet

Platelet

Vasc. Endo. Cell

PMN

ECM

Fibrinogen

1

2

3

2. Integrins bind to other cell adhesion molecules

2-Integrins promote platelet aggregation through soluble miltivalent mediator molecules

Integrin and platelet aggregation

(Karp, 2001)

Activation of some integrins may require to bind their ligand

Ex: Platelet aggregation

Leukocyte migration

Fibrinojen Trombin

thrombin activation of platelets causes an induction of platelet agregation mediated by fibrinogen binding to the integrin

Disintegrins

• Disintegrins are peptides isolated from the venom of various snakes of the viper family. They interact with the beta 1 and beta 3 families of integrin proteins.

• They cause the bleeding.

Disintegrin drugs

Drug Function

Bitistatin platelet aggregation inhibitor, which binds with high affinity to the alphaIIbbeta3 integrin

Kistrin Kistrin has an RGD site that competes for the platelet IIb/IIIa integrin

Barbourin that function as potent inhibitors of both platelet aggregation and integrin-dependent cell adhesion

Batroxostatin They were first identified as inhibitors of platelet aggregation and were subsequently shown to bind with high affinity to integrins and to block the interaction of integrins with RGD containing proteins for example they block the binding of the platelet integrin _IIb_3 to fibrinogen

Integrins and Signal Transduction:

• Integrins play an important role not only in structure & architecture of tissues, but also for signal transduction leading to regulation of functions in cell.

3-Immunoglobulin (Ig) Superfamily:

• Contain one or more Ig-like domains that are characteristic of antibody molecules

• a homophilic mechanism (between N-CAM molecules on adjacent cells). Some Ig-like cell-cell adhesion proteins, however, use a heterophilic mechanism

• Function: ICAM and VCAM molecules play an important role in T cell interactions and binding of leukocytes to activated or resting endothelial cells

•Mediate Ca2+-independent Cell-Cell Adhesion

3-Immunoglobulin (Ig) Superfamily : members

• Intercellular adhesion molecules (ICAMs)- on endothelial cells Vasculer adesion molecules: VCAM

• the neural cell adhesion molecule (N-CAM),

• Platelet-endotel..: PECAM

4- Selectins

• Integral proteins• Extracellular domain bind

specific carbohydrates on other cells

• Most commonly found on epithelial cells, used to mediate interactions with leukocytes (white blood cells/immune cells)

P-selectin glycoprotein ligand-1

4- Selectins

• 3 types:

- P-selectin- blood platelets,endothelial cells

-L-selectin- white blood cells

-E-selectin- endothelial cells

Selectins are calcium-dependent (C-type) lectins (carbohydrate binding proteins)

• L-selectin -– on lymphocytes (neutrophils)– binds specialized sulfated mucins (‘peripheral node

addressins’ or PNAd) made by high endothelial venules (HEV)

– Can be shed upon lymphocyte activation

• P-selectin - early role in entry to site of inflammation

– in Weibel-Palade bodies in endothelial cells and -granules of platelets

– translocates to membrane in response to thrombin, histamine, C5a, etc

– binds PSGL-1, a tyrosine sulfated mucin - on neutrophils, some effector T cells

• E-selectin - delayed role in entry to site of inflammation

– cytokine inducible on endothelial cells (especially cutaneous)

– binds carbohydrate ligand (sialyl-Lex) on neutrophil glycoproteins /glycolipids and cutaneous leukocyte

antigen (CLA) on effector T cells

L = C-type lectin domainE = EGF-like domainC = complement regulatory domain

Lymphocyte Extravasation

Selectins are involved in extravasation

Inflammatory signals activate endothelial cells making P-Selectin undergo exocytosis

P-Selectin on the surface of endothelial cells binds a specific carbohydrate ligand (Sialyl Lewis -x) on leukocytes

The leukocytes attach to the endothelial wall and roll slowly on it

PAF and integrins are then activated and the leukocytes start to extravasate

Selectins

Figure 19-19b Molecular Biology of the Cell (© Garland Science 2008)

functions of selectins

animation

Lökosit Endotel

1-K.H P-selektin

2- İntegrin ICAM

3-İntegrin ECM

Diseases of cell adhesion molecules

Integrin diseases-1

• Leukocyte adhesion deficiency

(LAD) type I: – defects in 2 integrin -> defective

neutrophil migration to inflammed skin, peritoneum; lymphocytes less affected due to continued use of

– LAD patients have recurrent bacterial infections

• Other types of LAD involve defects in expression of glycosyltransferases needed to make selectin ligands and defects in intracellular signaling molecules needed for chemokine-mediated integrin activation

• EX: LAD 2 results from a lack of sialyl LewisX (defect of carbohydrate fucosylation). Interaction with endothelial E-and P-selectins is impaired

Integrin diseases-2

• Glanzmann Thrombasthenia

• : is an inherited bleeding disorder – IIbß3 integrin

expression deficieny – ↓

• adequate formation of the platelet plug

» ↓

Susceptibility of bleeding

fibrinojentrombin

Immunoglobulins

• One Ig cell adhesion protein known as L1, assists in growth of nerve cells – mutants can have severe neural problems: – Mental retardation– Hydrocephaly (fluid

accumulation in ventricles of brain - arrows)

Cancer Metastasis and adhesionAlteration in expression of adhesion moleculescell- cell a recognition- adhesion system breaks down in cancer

6 basic steps for metastasis

For metastasis

1. DetachmentNo need for cell to cell interaction

2. Invasion and intravasation3. Circulation4. Stasis5. Extravasation and invasion6. proliferation

Cancer Metastasis and adhesion

cell- cell a recognition- adhesion system breaks down in cancer

• A loss-of-function allele of a Cadherin, a Desmocollin, or a Ca++-independent CAM can lead a cell to a lose its adhesion to proper neighbors and start to wander(tour)

• About 85% of cancers are epithelial in nature (due to high replication rate), metastasis is linked to a loss of E-cadherins

E (Cadherins is down regulated in most carcinomas and is therefore a tumor suppressor

E (Cadherins:

•.Loss of cadherin is accompanied by a loss of zonula adherens junctions and a dramatic reduction in cell-cell adhesion.

•Experimentally increasing the levels of E-cadherin can restore many of the normal •epithelial properties of carcinoma cells including loss of their ability to cause tumors• when injected into animals.

cadherins

CirculationStasis

Extravasation and invasion

Changes in cell adhesion behavior