Cell interactions

• Cellular interactions are required for?

  • intracellular communication
  • survival
  • tissue strength
  • organ function
  • immune system function
  • embryonic development
• Cadherins

  • Ca2+ dependent
  • used for adhesion or to transmit signals
  • binds to a similar cadherin on a neighbouring cell
  • Extremely important factor for molding cells into cohesive tissue in the embryo and holding them together in the adult
  • distributed along cell surfaces or part of intracellular junctions
  • synapses
  • adherens junctions
  • desmosomes
• 4 different families of integral membrane proteins mediate cell-cell adhesion. What are they?

  • Cadherins
  • Selectins
  • Integrins
  • IgSF (immunoglobin super family)
• Immunoglobulin super family (IgSF)


  • Contain Ig domains can connect to the integrin family, or connect to another IgSF
  • Mediate Ca2+ independent adhesions
  • Many IgSF proteins are ICAMs
  • ICAMs - intercellular adhesion molecules
  • Integrins are some of the proteins that acts as receptors for ICAMs
• What do we know? What are the basic cell interactions in neutrophil trapping?

  • Endothelial cell to Neutrophil: E-selectins to carbohydrate
  • Endothelial cell to Neutrophil: Platelet activating factor or IL-8 to G-protein coupled receptors (neutrophil)
  • Neutrophil to endothelial cell: Integrins (neutrophil) to ICAMs (endothelial)
• Cadherins mediate cell-cell recognition during embryogenesis

  Cells from different germ layers have different adhesive properties and the selective cell affinities help establish the spatial order of different tissues in the embryo.

  • The adhesive properties requires specific cadherin-cadherin interactions
• Selectins

  1. E-selectin, present on endothelial cells
  2. P-selectin, present on platelets and endothelial cells
  3. L-selectin, present on all types of leukocytes (white blood cells)

  • Ca2+ dependent
  • Selectins are glycoproteins that bind to specific oligosaccharide
  • Selectins have a small cytoplasmic segment, a membrane-spanning domain, and a large extracellular portion
• Neutrophils during inflammation

  1. Inflammation activates endothelial cells, which upregulates the selectins so they become more adhesive to the neutrophil
  2. Selectins bind to the carbohydrate residues (Psg-1) on neutrophils
  3. Platelet activating factor or IL-8 on the surface of endothelial cells activates G-protein coupled receptors on the neutrophil and this leads to integrin activation
  4. Integrins bind to ICAMs on endothelial surface and a cascade of events results in cytoskeletal rearrangement such that the cell can extravasate (leave the blood stream)
  5. Transendothelial migration
• Visual drawing by moi on neutrophil trapping

  A) Selectin

  B) GPCR

  C) integrin

  D) ICAMS

  E) IL-8

  
• One of the most important proteins that reduces metastasis is the presence of E-cadherins

  • Primary tumour = where cancer originates from. If high levels of E-cadherins then metastasis is unlikely
  • Secondary tumour = cancer has spread around the body
• The junctional complex

  • Tight junctions (zonula occluden)
  • Adherens junctions
  • Desmosomes (macula adherens)

  
• Tight junctions

  Found in the apical membrane. Occur between neighboring epithelial cells, prevent solute distribution where different solute concentrations are in adjacent compartments

  • TJ proteins such as claudins come from each cell and interact to form the junction
  • TJs can interact with actin and microtubules
• Polarity of epithelial cell

  A) Apical membrane

  B) Baso-lateral membrane

  
• Tight junction gate function

  
Controls the passage of ions, proteins, and water between the plasma membranes (paracellular pathway)
• Tight junction fence function

  Forms a barrier that blocks proteins to control the diffusion of integral membrane proteins between the apical and baso-lateral membrane of a cell, thus maintaining the cell's polarity.

  • Connects to the actin cytoskeleton and microtubules
• Adherins Junction

  • connects external environment to actin cytoskeleton
  • Provide a pathway for signals to be transmitted from the exterior to the cytoplasm and nucleus
  • form a belt (zonula adherens) that encircles the cells near their apical surface in epithelial cells
  • contain E-cadherins to connect the adherin to the two cells and is calcium dependent
• Desmosomes

  • Protein rich junctions
  • Contain cadherins that interact with multiple proteins to form a cytoplasmic plaque on the inner surface of the plasma membranes keeping the two adjacent cells together and anchoring the keratin intermediate filaments
  • the keratin intermediate filaments cytoskeleton anchors the the 2 cells together
  • provide strength to a sheet of cells
  • Part of the baso-lateral membrane

  
• Keratin IFs are in epethelial cells

  • form heterodimers (2 different keratins) and form long cables
  • contribute to the cell and tissue strength
  • Mutation in keratin genes can result in skin fragility
  • Gene expression and heterodimers change with specialization

  
• Keratin IFs are an important cytoskeleton in epithelial cells

  • keratin 5 and 14 are basal cell keratins and are the least specialized. They are the dividing cells and behave like stem or progenitors for the growing epidermis.
  • Basal layer marker
  • have a different cytoskeleton
  • keratin 1 and 10 are suprabasal cell keratins and are the most specialized. They are differentiated from keratin 5 and 14
  • They lose their nucleus (become cornified) and their proteins become crosslinked to prevent dehydration of the skin
• Gap junctions- communication channels


  • Form intercellular channels that transmit small molecules and ions directly through the membrane of adjacent cells and into adjacent cytoplasms
  • made of connexin proteins
  • 6 identical connexins from each cell form a transmembrane channel with a central pore called a connexon
  • 2 connexons form a gap junction
• Interactions involving the cell substratum

  • when cell wants to remain stationary: hemidesmosome
  • when cell wants to be motile: focal adhesions
• Hemidesmosomes

  • cell-matrix attachment to the basement membrane
  • A thick cytoplasmic plaque containing keratin filaments link the cell to the ECM
  • keratin filaments are linked to the ECM by integrins
• Focal adhesions

  • key role in cell locomotion
  • contain both actin and integrins
  • The attachment of integrins from a cell to the ECM activate focal adhesion kinase (FAK) and SRC which sends a survival signal to the nucleus
  • without activation of FAK and SRC, cells undergo apoptosis
  • Cells that have FAK and SRC always turned on can leave the ECM and migrate without undergoing apoptosis
• Model of integrin activation


  • binding of protein, talin, to the beta subunit of integrin induces a separation of the alpha and beta subunits and conversion to an active conformation.
  • Active conformation has subunits that are extended and separated
  • inactive conformation has a bent structure
  • Talin connects to actin and helps transmit the signal to help it move
• Inside-out activation: signal activating the integrin comes from within the same cell. Eg. neutrophil trapping; signal came from GCPR on neutrophil

  outside-in activation: binding of a substrate not of the same cell activates integrin
• How do some integrins contact other proteins?

  Integrins have a RGD binding site that binds to a RGD peptide in its ligand.

  • not all integrins have RGD binding sites (only about half)
• If an enzyme that remove hyaluronic acid was added to extra-cellular matrix proteins, which proteins would be affected?

  proteoglycans
• What data led researchers to conclude that the signaling activity resulting in cAMP production in fat cells was due to multiple receptors types activating a shared pool of adenylyl cyclase enzymes?

  Upon treatment with multiple hormones, the cAMP response was not additive when compared to treatment with individual hormones
• Signaling enzymes will dock to the RTK if they have SH2 domains and are often activated by this association.
• STAT5 is in the cytoplasm during activation but will translocate to nucleus after activation to help transcribe genes
• Stats can be shut off by removing the phosphoryl group from the tyrosine or the presence of proteins that bind to the receptor and prevent docking to the receptor
• The steps of a generalized MAP kinase cascade

  • Recruitment of Grb-Sos (GEF factor) after receptor activation. Grb2 is an adaptor protein with an SH2 domain to bind to receptor.
  • Sos exchanges GDP for GTP in Ras proteins to activate them. Ras recruits Raf that will activate membrane-bound Ras (MAPKKK) which will start the MAPK phosphorylation cascade.
  • Once ERKs (MAPK) are phosphorylates (active) they activate TFs Jun and Fos to activate transcription of gene cd1 (promote cell cycle) and a later gene MKP-1 that removes phosphoryl group from MAPK
• • Convergence: Signals from unrelated receptors lead to activation of a common effector (eg. different hormones that activate adenyl cyclase)
  • Divergence: A signal reaches a variety of effectors. (eg. signal transduction)
  • Cross-talk: Signals can be passed back and forth between pathways and influence each other's pathways. Convergence and divergence are examples of cross talk.
• convergence of signals of MAPK pathway

  GPCRs, receptor tyrosine kinases, and integrins bind to different ligands but they all can lead to a docking site for Gbr2
• Steroid hormones can enter the cell and bind their receptor in the cytoplasm

  • Cytoplasmic receptors can directly control gene expression by traveling to the nucleus and binding DNA, recruiting coactivators or corepressors
• Steroid nuclear hormones and receptors

  Eg. glucocorticoids, progestins, estrogens, androgens, and mineralocorticoids

  • Type I receptors, androgen receptor, the estrogen receptor, and the progesterone receptor, are present in the cytoplasm bound to chaperone proteins (e.g., HSP90)
  • Ligand binding results in release of the receptor from the chaperone, homodimerization, and translocation of receptor into the nucleus.
• Steroid hormone receptors are receptors that are transcription factors


  TF domains:

  • DNA binding domain: binds to specific DNA sequences in a gene promoter
  • Trans-activation domain: activates transcription
  • ligand binding domain (unique to hormone receptors): bind to the nuclear receptor to activate the receptor, which can result in nuclear translocation of hormone receptor
• What happens after steroid hormone receptor is translocated?

  The active receptor complex binds the DNA response element and associates with transcriptional coactivators that help activate target genes. Can also recruit corepressors
• The glucocorticoid receptor

  • The glucocorticoid receptor (GR) is a nuclear receptor that includes a ligand-binding domain and a DNA-binding transcription factor.
  • The GR binds to a glucocorticoid response element (GRE), which is a palindrome (when two DNA strands have the same 5' to 3' sequence)
  • GR is a zinc-finger protein: Fingers independently project into successive major grooves in the target DNA
• How is space created for all the transcription factors to fit on the gene promoter?

  TFs can modify the chromatin state:

  • DNA methylation
  • Histone acetylation
  • Histone methylation