2. Cell Adhesion and Communication

Learning Outcomes

• Describe with examples the structure and function of different classes of cell-cell and cell-matrix interactions
• Give an overview of how cell adhesion molecules can interact with components of the cytoskeleton
• Give an overview of the role integrins play in cell signalling events

Epithelial Tissue

• Covers the internal and external surface of organs
• Contains a layer(s) of cells on top of basement membrane
• Cells adhere to each other laterally as well as to basement membrane
• 

Cell adhesion overview

• Lateral Cell-Cell Adhesion established via:
• Tight junctions (junctions are specialised modifications of cell surface in neighbouring cells)
• Adherens junctions
• Desmosomes
• Gap junctions
• Non-junctional adhesions (proteins such as cadherins, selectins, integrins)
• Cell-Matrix Adhesion established via:
• Hemidesmosomes
• Focal adhesions
• Non-junctional adhesions (integrins)
• 3 distinct classes of cell adhesion junctions:
• Occluding (tight) junctions= forms a barrier between epithelial cells
• Attachment junctions= links cell cytoskeleton to that of its neighbouring cell or to matrix
• Communication junctions= directly connect cytoplasm of adjacent cells

Tight junctions: Structure

• Structure: Proteins
• Seal adjacent epithelial cells in a narrow band beneath their apical surface
• Bands made up of interconnected (sealing) strands of transmembrane proteins that wrap around the cell
• 3 types of transmembrane proteins:
• Claudins = core proteins in tight junction fibrils
• Occludins = position and stabilise claudins
• Junctional adhesion molecules (JAMs) = position and stabilise claudins
• Proteins in the sealing strands attach stably to structural proteins to interconnect the sealing strands, also transiently to signalling proteins
• 
• Bildunterschrift: Electron micrograph- network of thin fibrils where protein embedded (left) 2 plasma membranes of adjacent cells joined together by tight junctions (right)

Tight junctions: Selectivity

• Tight junctions allow the passage of small ions, solutes, electric currents between epithelial layers
• Differing junctions allow passage of differing solutes through
• Dependent on size (4-40A) and charge, different tissues have different selectivities)
• Selectivity determined from different claudins- humans have 24
• Eg expression of claudin 16 restricted to ascending limb of loop of Henle, mutations associated with conditions caused by Mg and Ca ions unable to pass through tight junctions

TJ: Role- regulate paracellular transport

• 
• Transfer substances across epithelium by passing through intracellular space between cells
• Act as molecular sieves rather than completely impermeable ("occluding")
• Property of TJs which regulates this process= selective permeability

TJ: Role- regulate transcellular transport

• 
• Transfer substances across epithelium by passing through cells, crossing apical and basolateral membranes
• Allows substances against concentration gradients eg Glu absorption
• Property of TJs which regulates this process=separation of apical and basolateral membrane

Transcellular transport: Glucose absorption

• If absorption of glucose across intestinal epithelium relied on paracellular transport we would not gain all the glucose from our diet:
• Sodium-potassium pump ATPase in basolateral membrane generates Na gradient by pumping Na out of the cell against conc gradient
• Low conc of Na in cell compared to outside
• Energy stored in gradient is used to bring glucose into cell from lumen of SI via Na/Glu symporter
• High conc Glu in cell compared to bloodstream
• Glu enters bloodstream via facilitated diffusion through Glu trasnporter (GLUT2 in baolateral membrane)
• Tight junctions play a role in maintaining separate identity of 2 membranes and membrane proteins cant cross the barrier they create:
• Essential that ATPase and GLUT2 are present on basolateral membrane and not apical
• Essential that Na/Glu symporter is on apical and not basololateral

Tight Junctions: Summary

• Formed by interconnected sealing strands of claudin, occludin and junctional adhesion molecules that encircle the cell
• Act as molecular filters thus regulating paracellular transport across epithelial tissue
• Maintain identity of apical and basolateral membranes by preventing diffusion of proteins and lipids between the two

Attachment Junctions: Adherens

• 
• Bildunterschrift: Electron micrograph- thick, dark bands close to PM of neighbouring cells, bridged by rod-like structures projecting into extracellular space
• Serve as bridges: link actin filaments (through catenin) of adjacent cells
• Zonula adherens: found immediately beneath tight junctions in epithelial tissues
• Other types found joining cell types

Folie 11

• All adherens junctions have 2 common properties:
• Contain cadherins:
• TM proteins that bind to identical cadherins on neighbouring cells
• Role in determining tissue organisation- enable cells to find appropriate binding partner and determine junction strength
• Link to actin cytoskeleton:
• Regulate changes in cell shape and/or to sheer stress (important in cytoskeleton dynamics)
• Through catenins (role in signalling : beta-catenin can act as TF)
• Catenins link cadherins to MFs eg link cytoplasmic tails of cadherins to MF in zonula adherens, mutations in catenins lead to cancer

Attachment junctions: Desmosomes

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• Bildunterschrift: Electron micrograph- thick patches of electron-dense material on cytoplasmic side of PM, connected to int filaments in cytosol
• Also add structural integrity to tissues by linking cytoskeleton of adjacent cells
• Through intermediate filaments

Attachment junctions: Desmosomes

• 2 common properties:
• Contain cadherins (desmocollins and desmogleins) that link adj cells together:
• Cadherins bridge intracellular space on one side of membrane and serve as docking sites for cytosolic proteins (desmoplakin) that bind int filaments on the other side
• Link to int filaments through adaptor proteins( plagoglobulins/desmoplakin):
• Attachment to filaments through adaptors regulated by cell signalling
• Can be initiated by activated growth factor for receptors (coordination of cell growth and adhesion)

Attachment junctions: Desmosomes

• 
• Electron micrograph of desmosome between 2 keratinocytes:
• Shows intermediate filaments linking to cytoplasmic plakoglobulin and desmoplakin that, in turn link to TM cadherins desmoglein and desmocolin
• Formation of bridge between plasma membrane of neighbouring cells

Attachment junctions: Hemidesmosomes

• Located on basal surface of epithelial cells
• Providing structural stability to epithelial sheets
• Anchor cells to underlying basement membrane through cytoplasmic 'plaques' that connect to intermediate filaments
• Plaques contain integrins (TM protein) which connect to int filaments through adaptor protein plectin
• Extracellular region of integrins binds to components of basement membrane

Arrangement of hemi- and desmosomes

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• Organises intermediate filaments into robust network
• Mutations that disrupt desmosomes and/or hemidesmosomes -> severe blistering diseases
• Hemi and desmosomes both link intermediate filaments but are structurally very different

Integrins: general

• Heterodimers receptors composed of alpha and beta subunits: 18 different alpha, 8 beta, 24 known different heterodimers
• Bind to ECM components:
• Different heteromers bind different ECM components- different cell types express different integrins so binding of a cell to its ECM is determines by the integrins present on its basolateral membrane
• Specificity of which ECM component an integrin binds is determined by integrin alpha subunits
• Complexity: Most heterodimers can bind more than one ECM component and each component binds more than one integrin
• All known binding sites on ECM proteins contain acidic amino acid (asp) and many contain sequence : Arg-Gly-Asp
• Also bind to components of focal adhesions- linking actin cytoskeleton to ECM

Integrins: Signalling

• Changes within the cell affect affinity of the integrin for its ligands
• Activation of adaptor proteins (eg talins) -> link integrins to actin cytoskelton focal adhesions:
• Integrin doesnt bind to actin cytoskleton or ECM until talin binds
• Signalling via GRCP or RTK activates talin and induces dimerisation
• Talin binds to beta-intgrin subunit which causes conformational change in integrin extracellular binding domain, enabling ECM binding
• On binding, talin also links actin filaments
• Inside-out signalling: signalling pathways inside cell affect role of integrins outside
• 

Integrins: Signalling

• Outside- in signalling: transduce information outside of the cell to the inside
• Binding of cell to underlying matrix can cluster integrins together
• Makes their cytosolic domains form a docking site for signalling molecules
• Integrin signalling pathways regulate many cell physiologies:
• Cell differentiation
• Inhibition of programmed cell death
• Cytoskeletal rearrangements: importance in cell migration across the ECM and not sticking to it eg during wound-healing focal adhesions promote remodelling of actin cytoskeleton to drive leading edge forward

Trafficking of integrins

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• Trafficking of integrins used to regulate cell movement: formation/disassembly of focal adhesions
• Endocytosis of integrins:
• Can break attachment to substrate
• Internalised integrin can be recycled and establish new attachments in direction of movement
• Or integrin can be trafficked to the lysosome and degraded causing the cell to loose attachment = dysregulation in cancers

Communication junctions (Gap)

• Gap junctions facilitate direct transfer of ions and small molecules between adjacent cells
• Found in most vertebrate and invertebrate cells and contain upto thousands of channels- can form large or small junctions
• Only known means of direct cell-to-cell transport in animals
• 
• Bildunterschrift: Regions where PMs are closer together than surrounding areas with gaps bridges by channels projecting out of PM

Gap junctions

• Gap junction channels consist of 2 halves (hemichannel or connexons):
• 2 connexons on opposing membranes of adjacent cells dock togther to form gap junction
• In intracellular gap creates 2-4nm pore for ions/small molecules to pass through
• Subunit of connexon= connexin
• 6 connexins form hetrameric connexon = channel cluster
• Connexin subunit = tetraspan, 4 TM alpha helices
• Gap junction channels have different permeability depending on connexins present: Homomeric/homotypic, heteromeric, heterotypic
• Transport: inorganic ions, sugars, amino acids, nucleotides, vitamins, cAMP and IP3, NOT macromolecules (proteins, NA, polysaccharides)

Gap junctions: Function and Regulation

• Function: Used for rapid communication
• Reflexes reactions: in brain mediated by neurons linking gap junctions, allowing potential to be spread rapidly, avoiding delay at chemical synapses
• Synchronisation of cardiomycetes signal to contract: communicated through gap junction, mutations in connexins lead to arrythmia
• Regulation
• Alternate between open and closed states
• Closed- through phosphorylation of connexins when large increase in intracellular Ca ion conc and/or pH
• Eg cell damage causes elevation of Ca in cell and releases damaging metabolites
• Closing prevents further damage to neighbouring cells

Summary

• Connections between cells can be established via:
• Tight junctions
• Adherens junctions
• Desmosomes
• Gap Junctions
• Synapses
• Connections between cell & their underlying matrix can be established via:
• Hemidesmosomes
• Focal adhesions
• Non junctional adhesions