MCB 252 - Topic 13 - Integrating Cells into Tissues Part 1: Cadherins and Cell Adhesion

Types of Tissues

  • Four major types of tissues:
    • Epithelial Tissue
    • Connective Tissue
    • Muscle Tissue
    • Nervous Tissue

Epithelial Tissue

  • Epithelia:
    • Line or cover surfaces or body cavities
    • Functions include:
      • Exchange
      • Secretion
      • Absorption
      • Protection
      • Waterproofing
  • Types of Epithelial Tissue:
    • Simple squamous epithelium
      • Location: Air sacs of lungs and the lining of the heart, blood vessels, and lymphatic vessels
      • Function: Allows materials to pass through by diffusion and filtration, and secretes lubricating substance
    • Simple cuboidal epithelium
      • Location: In ducts and secretory portions of small glands and in kidney tubules
      • Function: Secretes and absorbs
    • Simple columnar epithelium
      • Location: Ciliated tissues are in bronchi, uterine tubes, and uterus; smooth (nonciliated tissues) are in the digestive tract, bladder
      • Function: Absorbs; it also secretes mucous and enzymes
    • Pseudostratified columnar epithelium
      • Location: Ciliated tissue lines the trachea and much of the upper respiratory tract
      • Function: Secretes mucus; ciliated tissue moves mucus
    • Stratified squamous epithelium
      • Location: Lines the esophagus, mouth, and vagina
      • Function: Protects against abrasion
    • Stratified cuboidal epithelium
      • Location: Sweat glands, salivary glands, and the mammary glands
      • Function: Protective tissue
    • Stratified columnar epithelium
      • Location: The male urethra and the ducts of some glands
      • Function: Secretes and protects
    • Transitional epithelium
      • Location: Lines the bladder, uretha, and the ureters
      • Function: Allows the urinary organs to expand and stretch

Connective Tissue

  • Examples:
    • Loose connective tissue
      • Locations: Beneath dermis of skin, digestive tract, respiratory and urinary tracts; between muscles; around blood vessels, nerves, and around joints
      • Functions: Cushions organs; provides support but permits independent movement; phagocytic cells provide defense against pathogens
    • Adipose tissue
      • Locations: Deep to the skin, especially at sides, buttocks, breasts; padding around eyes and kidneys
      • Functions: Provides padding and cushions shocks; insulates (reduces heat loss); stores energy reserves
    • Dense connective tissues
      • Locations: Between skeletal muscles and skeleton (tendons); between bones (ligaments); covering skeletal muscles; capsules of internal organs
      • Functions: Provides firm attachment; conducts pull of muscles; reduces friction between muscles; stabilizes relative positions of bones; helps prevent overexpansion of organs (such as the urinary bladder)
  • Connective Tissue Types:
    • Loose connective tissue
      • Cells: Fibroblasts, adipose and white blood cells
      • Matrix Composition: Loose elastin and collagen networks
      • Cells to Matrix Site: High, Under skin
      • Function: Anchor, Support
    • Dense connective tissue
      • Cells: Fibroblasts
      • Matrix Composition: Dense elastin and collagen networks
      • Cells to Matrix Site: Low, Ligaments and tendons
    • Adipose tissue
      • Cells: Adipocytes
      • Matrix Composition: Cells with abundant lipid
      • Cells to Matrix Site: High, Beneath skin, between muscles, around heart and joints
      • Function: Padding/protection, Insulation
    • Blood
      • Cells: Red and white blood cells, platelets
      • Matrix Composition: Plasma
      • Cells to Matrix Site: High, Throughout the body
    • Cartilage
      • Cells: Chondrocytes
      • Matrix Composition: Collagen
      • Cells to Matrix Site: Low, Ears, joints, bone ends, respiratory passages, embryonic skeleton
      • Function: Connect, Cushion
    • Bone
      • Cells: Osteoclasts, osteoblasts, osteocytes, osteoprogenitor cells
      • Matrix Composition: Collagen, minerals
      • Cells to Matrix Site: Low, Skeleton

Muscle Tissue

  • Three types of muscle tissue:
    • Skeletal Muscle Tissue
      • Function: Skeletal muscles move or stabilize the position of the skeleton; guard entrances and exits to the digestive, respiratory, and urinary tracts; generate heat; and protect internal organs. Skeletal muscle tissue contractions move the body by pulling on bones of the skeleton, making it possible for us to walk, dance, bite an apple, or play the ukulele.
    • Cardiac Muscle Tissue
      • Function: Cardiac muscle moves blood and maintains blood pressure. Cardiac muscle tissue contractions in the heart propel blood through the blood vessels.
    • Smooth Muscle Tissue
      • Function: Smooth muscle moves food, urine, and reproductive tract secretions; controls diameter of respiratory passageways and regulates diameter of blood vessels. Smooth muscle tissue contractions move fluids and solids along the digestive tract and regulate the diameters of small arteries, among other functions.

Neural Tissue

  • Composed of:
    • Sensory Neuron
    • Motor Neuron
    • Interneuron
    • Plus glial cells and other specialized cells

Evolution of Multicellular Organisms

  • Some Major Transitions in Evolution
    • 3. 5 billion years ago: unicellular life emerges; photosynthetic bacteria begin to release oxygen
    • Eukaryotes emerge – longer ago than 2.7 BYA
    • 555 million years ago; Multicellular organisms common in the sea. Single cell organisms -> multicellular organisms predates this time
    • 500 million years ago vertebrate fish
    • 450 million years ago Arthropods move onto land, descendants become spiders, mites, scorpions and millipedes
    • 420 MYA Land plants evolve/emerge
    • 360 MYA 4-limbed vertebrates move onto land as seed plants and large forests emerge
  • “Philosophical” or life-style differences between unicellular organisms and multi- cellular organisms
  • C. elegans: Development follows an “Invariant” Lineage
    • Apoptosis- one unique feature in multicellular organisms
    • 1090 Cells formed
    • 131 Undergo “Programmed Cell Death”/Apoptosis = 12%

Cell Adhesion Experiments

  • Early experiments to understand cell adhesion
    • H.V. Wilson (1907)- sponge cell adhesion
      • Mechanical dissociation of sponge cells -> individual cells reaggregate
  • Experiment using two different species:
    • Mechanical dissociation -> mix -> sort
  • Embryogenesis
    • Red = ectoderm
    • Blue = Neural
    • Green = mesoderm
    • Yellow = endoderm
  • Reaggregation of amphibian embryonic cells (Townes and Holtfreter, 1955)
    • Dissociation of cells -> Spontaneous reaggregation -> Segregation of cell types
  • Sorting in Amphibian Embryonic Cell Aggregates
    • Examples:
      • Epidermis + Mesoderm
      • Mesoderm + Endoderm
      • Neural plate + Epidermis
      • Neural plate + Mesoderm + Epidermis
      • Epidermis + Mesoderm + Endoderm
  • Cell Sorting in Embryonic Organs
    • Dissociate embryonic organs -> Combine -> Cells reaggregate -> Sort according to organ type -> Specific patterns of aggregation
    • Differential adhesion hypothesis (Steinberg): Cells sort due to differences in adhesive properties (could be quantitative or qualitative)

Discovery of Cadherins

  • How Can One Identify the Hypothetical Adhesion Molecules?
    • First, need an assay: in vitro aggregation.
      • Dissociated cells in Petri dish
    • Where are the molecules that mediate adhesion localized?
      • Look for antibodies against surface proteins that can block aggregation
  • Adhesion blocking antibodies were used to identify the Ca2+-dependent adhesion molecules
    • Screen cDNA expression library
  • How would you “identify” a protein (a band on a gel) in the genomic era?
    • Immuno-precipitation Experiment
      • Extract + Antibody + Bead with Protein A attached -> Add bead and antibody -> Ppt bead, antibody and your protein -> Remove supernatant -> Resuspend your protein in buffer
    • Use antibody to immuno- precipitate the protein of interest or to identify which band on a gel is your protein.
    • The cDNA encodes a putative cell-adhesion molecules (CAM); how do you convince yourself that it really is a CAM?
      • Identify protein based on protein composition and mass compared to a data based of information on each and every protein in the genome.
  • Expressing Cadherins in a Fibroblast Confers Cell-Cell Adhesive Properties.
    • Fibroblasts and certain cancer cells show very little cell-cell adhesion even in the presence of Ca2+Ca^{2+}
    • Transfecting these cells with a cDNA encoding for E-cadherin confers the property of calcium dependent cell-cell adhesion.
  • Discovery of Cadherin
    • Takeichi, J. Cell Biol. 1977:
      • Cell aggregation in calcium
      • + non-immune control Ab.
      • + novel antibody that disrupts cell-cell adhesion.
    • Yoshida and Takeichi, 1982
      • Antibody recognizes a band on cells in the presence of calcium but not in the absence of calcium.
  • An RNAi Experiment Demonstrating that Cadherins are Necessary for Strong Cell-Cell Adhesion.
    • Heasman and Wylie.
      • Cells inside this embryo which has been depleted of the major cadherin expressed at this time (C-cadherin) detach from one another.
      • Cells inside this normal frog embryo are closely packed together.
  • Early embryology experiments showed that the different cell types selectively adhere to one another.
  • E-cadherin confers cell-cell adhesive properties to a non-adherent cell.
  • Might animals express multiple cadherins to help drive tissue sorting?
  • Different tissue types segregate from one another during development.
    • Separation of the neural tube from the overlying ectoderm is a classic case.

Cadherin Family

  • Cadherins Comprise a Family of Adhesion Molecules with Differential Tissue Distribution.
    • The ecotoderm expresses E-cadherin
    • The neural tube expresses N-cadherin
  • Expressing Two Different Cadherins is Sufficient to Drive Cell Sorting
    • Express P-cadherin in one population and label them green.
    • Express E-cadherin in another and label them red.
    • Mix and add calcium.
    • Populations sort out over time.
      • Later time points – E sorts away from P.
      • Early time points – E and P intermingle.
  • The Cadherin Superfamily
    • Cadherin superfamily is characterized by a cadherin motif called an EC domain.
      • Based upon this, humans express at least 80 different cadherin superfamily proteins.
      • Are they all adhesion molecules?
      • Why this complexity?
    • First cadherins discovered were the “classic cadherin” and these have a conserved domain structure.
      • Classic cadherins mediate cell-cell adhesion.
    • Research on the other types of cadherin family members is just getting started.
      • Thus far, the few protocadherins that have been studied in detail are anti-adhesive (!)
  • Differential Cadherin Expression Demarcates Regions of the Developing Brain.
    • Takeichi and others think that the diversity of cadherin family members might help explain complex segregation of different cell types.
    • Different cadherins, for example, are expressed in discrete regions of the brain.

Cadherin Structure and Function

  • Model of Cadherin structure
    • 5 extracellular repeated domains (Cadherin repeats)
    • transmembrane (TM) domain
    • conserved intracellular domain
  • Models of Cadherin structure
    • 5 extracellular repeated domains (Cadherin repeats)
    • transmembrane (TM) domain
    • conserved intracellular domain
    • role of Ca2+Ca^{2+}
  • How is Cadherin binding specificity determined?
    • Domain swapping experiments
      • Exchange different portions of the extra-cellular repeats, assay binding mix with cells expressing either P- or E-cadherin (labeled red)
      • N-terminal 113 aa essential for specificities (1st cadherin repeat)