22 August 2006
Cell Structure and Function II
Dr. Joana Chakraborty, Ph.D.



Cytoskeleton


  • Actin dynamics for membrane extension
    • Can be in globular or filament form
  • Filament networks (bundles) for maintaining cell structure
  • Myosin motors for contractility and vesicle transport
  • Actin bundles and intermediate filaments for cell adhesion
  • Lamin network (intermediate filament) for nuclear structure
    • Inside of nuclear envelope
  • Microtubules work with other cytoskeleton components for proper function

Actin Filaments and Microfilaments

  • Structure
    • Thinnest filament, ~6 nm
    • 15% of total protein of a cell
    • G-actin and F-actin types
    • Actin filament has a + and a – end and is dynamic – forming when needed and dissociating when not needed
      • Most material joins on the + end when forming and most material leaves on the – end when dissociating
      • Actin grows fast on + end and very slowly on - end
    • Three classes of actin – α in muscle cells and , β and γ in non-muscle cells
  • Actin-Binding Proteins
    • Necessary for proper function – defects can lead to disease
    • Fimbrin
      • Microvilli, stereocilia, adhesion plaques, yeast actin cables
      • Help actin filaments bind side-by-side to form bundle
    • α- Actinin
      • Filopodia (extension of cell membrane), lamellipodia, stress fibers, adhesion plaques
      • Help actin filaments bind side-by-side to form bundle
    • Spectrin
      • Cortical networks
      • Allows cell to retain flexibility necessary for cells such as blood cells to move through the thin corridors of capillary systems
    • Dystrophin
      • Muscle cortical networks
      • Muscular dystrophy occurs when dystrophin is defective
    • Filamin
      • Filopodia, pseudopodia, stress fibers
    • Fascin
      • Filopodia, lamellipodia, stress fibers, microvilli, acrosomal process
    • Villin
      • Microvilli in intestinal and kidney brush border
    • Cofilin
      • Dissociation from – end
    • Severin and Gelsolin
      • Severing, capping + end
  • Mechanism of Assembly and Disassembly
    • Globular actin molecules form a dimer, then a trimer, and continue to polymerize to form the helical actin filament through the nucleation process
    • Rapidly growing and slow growing ends
    • G-actin, Ca2+ and ATP needed
  • Actin Bundles and Filaments – formed by cross-linking proteins
    • Contractile, i.e. cleavage furrow (myosin) – can be used to push things around
    • Gel-like network, i.e. cell cortex (filamin)
    • Parallel, i.e. core of microvilli (villin, fimbrin)
    • Focal contact for cell attachment, i.e. stress fiber (α-actin, talin, vinculin)
  • Microvilli
    • Finger-like projections of surface of intestinal epithelium with variable length (0.5-10 μm)
    • Brush border of intestinal epithelium – increases surface area to transport nutrients by extending plasma membrane using actin filaments
    • Contains core actin filaments, membrane linkage, cross-liked by fimbrin and villin
  • Action of Drugs
    • Cytochalasins – used to prevent further polymerization of actin by binding to the ends of actin filaments
      • From various fungi
    • Phalloidin – binds to actin filaments and stabilizes against depolymerization
      • From mushrooms, very potent poison

Intermediate Filaments

  • Provide a strong 3-D structural support of cell
    • Kind of like bones of a cell
    • Anchor the nucleus in place
    • Connect the cell membrane and cytoskeleton
    • Form the framework for the maintenance of the nuclear envelope and nuclear structure
  • Structure
    • Two of them join to form a dimer, then two dimmers form a protofilament, elongate, and wrap up into an intermediate filament
  • Principle types of intermediate filament proteins
    • Cytoplasmic Intermediate Filament – can be used to identify the source of cancer primaries
      • Keratins – in epithelial, hair, nails
      • Desmin – in muscle cells
      • Vimentin – in fibroblast and leukocytes
      • Glial Fibrillary Acidic Protein – in astrocytes, Schwann cells
      • Neurofilaments – neurons
    • Nuclear Intermediate Filament
      • Nuclear Lamins – all nucleated cells
        • Control assembly of nuclear envelop by organizing perinuclear chromatin
        • Type B bines with inner nuclear member
        • Type A and C bind with B
  • Functions
    • Very Stable, distribute tensile strength across cell and tissue
    • No polarity (+ end or – end) because they do not assemble or disassemble easily

Microtubules

  • Structure
    • Has a + and – end polarity
  • Types of Microtubules
    • General Cytoplasmic
    • Central pair of cilia and flagella
    • Fused
  • Microtubule Associated Proteins – MAPs
    • Can organized microtubules and affect their stability
  • Assembly and Disassembly of Microtubules
    • Hollow tube composed of 13 protofilaments made from tubulin molecules form to make a microtubule
    • Can be fused in doublets (cilia, flagella) or triplets (basal bodies, centrioles)
    • Have a rapid growing end and slow growing ends
    • Mg2+ and GTP are needed
    • Some MAPs are needed also
    • Formed by α and β tubulin molecules into protofilaments arranged in a cylinder
  • Motor Proteins
    • Kinesin – Transport vesicles to + end
    • Dynein – Transport vesicles to – end
      • Present in cilia and flagella
  • Action of Drugs
    • Colchicine, Colcemid – inhibit addition of tubulin molecules to microtubules, leading to microtubule depolymerization
    • Vinblastine, Cincristine – induce the formation of paracrystalline aggregates of tubulin
    • Paclitaxel – Stablizes tubulin

Nucleus


  • Bound by nuclear envelope, contains nucleolus, chromatin, and nucleoplasm
    • Varying different shapes and sizes
  • Nucleus is the information center of the cell
  • The Interphase Nucleus
    • Distinct nuclear envelope, chromatin (light, filamentous material), nucleoplasm, and some centrosome and centriole (microtubules) structures outside of the nucleus
  • Nuclear Envelope
    • Double lipid bilayer with pore-complexes in envelope
    • Continuous outer membrane with rough ER
    • Inner membrane has nuclear lamin (30-100 nm)
  • Nuclear Pore Complex (NPC)
    • 80-100 nm, composed of proteins called nucleoporins
    • Necessary for precise transport of specific proteins in and out of nucleus
      • Larger molecules must interact with other proteins with nucleoporins for transport
    • Has 3 rings: cytoplasmic, middle, and nucleoplasmic
    • Cytoplasmic and middle rings are connected by 8 spoke-like structures
    • Nucleoplasmic ring has a protruding nuclear basket
    • Middle ring contains a central transporter
      • FG-nucleoporins line the central transporter channel and transport all macromolecules
    • Exportins – transport macromolecules to cytoplasm
    • Importins – transport proteins to nucleus
  • Nucleolus -- ribosome production factory
    • Fibrillar center of inactive DNA
    • Pars fibrosa – RNA
    • Pars granulose – maturing ribosomal units
    • Nucleolar matrix – has some nucleolus organization proteins
  • Chromatin – made up of DNA
    • Hematochromatin
      • Closer to nuclear envelope and is very dense
      • Not transcriptionally active
      • Low protein synthesis
    • Euchromatin
      • Much lighter than hematochromatin
      • Transcriptionally active
      • Meaning euchromatin cells are producing more protein than hematochromatin
  • Centrosome and Centriole
    • Centrosome
      • Microtubular organizing center (MTOC)
      • Some cells may not have centriole but will have centrosome instead
      • Centrosome is composed of the centrioles and surrounding pericentriolar material consisting of γ-tubulin ring complex, pericentrin, and other microtubule-nucleating macromolecules
      • Centrosomes assist in the formation and organization of macrotubules and self-duplication before cell division
    • Centriole
      • Centrioles in the normal in interphase cell are found in pairs at right angles to each other
        • Centrioles are composed of nine triplets of microtubules arranged around a central axis.
      • Centrioles when replicating will separate and bind with tubulin and begin forming microtubules at a right angle
      • Centrioles function in the formation f the centrosome and guide the formation of the spindle apparatus, cilia, and flagella
    • Microtubules (spindle fibers)
      • Kinetochore microtubules – connecting from centrosome to chromosome (“captures” chromosomes)
        • The location where the kinetochore microtubule attaches to the chromosome is called the kinetochore
      • Polar microtubules – from centrosome pole to pole
      • Astral microtubules – from centrosome and extend outwards
      • Microtubules grow outward from the poles

Mitotic Chromosome Movement

  • Kinetochore microtubules shorten from – end
  • Motor proteins on polar microtubules push them apart as polar microtubules elongate from + end
  • Astral microtubules shorten from – end while motor proteins also help pull
  • All three actions help separate chromosomes in anaphase
  • The assembly and disassembly of microtubules is what allows the movement of chromosomes (an any other organelles in the cell)
    • Control of assembly and disassembly controlled by availability of Mg2+ and GTP

Objectives