Fundamentals of Cell Biology

1
Fundamentals of Cell Biology

• Chapter 5 The Cytoskeleton and Cellular
  Architecture

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Chapter Summary The Big Picture (1)

• Chapter foci
• Cytoskeletal proteins form a skeleton inside the
  cell
• Intermediate filaments provide the cell with
  mechanical strength
• Microtubules are associated with cellular
  trafficking
• Actin is responsible for large-scale movements
• Eukaryotic cytoskeletal proteins evolved from
  early prokaryotes

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Chapter Summary The Big Picture (2)

• Section topics
• The cytoskeleton is represented by three
  functional classes of proteins
• Intermediate filaments are the strongest, most
  stable elements of the cytoskeleton
• Microtubules organize movement inside a cell
• Actin filaments control the movement of cells
• Eukaryotic cytoskeletal proteins arose from
  prokaryotic ancestors

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The cytoskeleton is represented by three
functional classes of proteins

• Key Concepts
• The cytoskeleton is a complex mixture of 3
  different types of proteins that are responsible
  for providing mechanical strength to cells and
  supporting movement of cellular contents.
• The most visible form of cytoskeletal proteins
  are long filaments found in the cytosol, but
  these proteins also form smaller shapes that are
  equally important for cellular function.
• The structural differences between the 3 protein
  types underscores their 4 different functions in
  cells.

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Cytoskeleton

• occupies large portion of cytosol and appears to
  link organelles to each other and to plasma
  membrane
• 3 elements IFs
• MTs
• Actin
• Elements do not form mixed polymers

Figure 05.01 The cytoskeleton forms an
interconnected network of filaments in the
cytosol of animal cells.
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IFs are the strongest, most stable elements of
the cytoskeleton

• Key Concepts (1)
• Intermediate filaments are highly stable polymers
  that have great mechanical strength.
• Intermediate filament polymers are composed of
  tetramers of individual intermediate filament
  proteins.
• Several different genes encode intermediate
  filament proteins, and their expression is often
  cell- and tissue-specific.

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IFs are the strongest, most stable elements of
the cytoskeleton

• Key Concepts (2)
• Intermediate filament assembly and disassembly
  are controlled by posttranslational modification
  of individual intermediate filament proteins.
• Specialized intermediate-filament-containing
  structures protect the nucleus, support strong
  adhesion by epithelial cells, and provide muscle
  cells with great mechanical strength.

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IFs provide mechanical strength to cells
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IFs are formed from a family of related proteins
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The primary building block of IFs is a
filamentous subunit

• a-helices in the central rod domain

Figure 05.04 The central rod domain of
intermediate filament proteins forms an alpha
helix. The head (and tail) regions form globular
shapes.
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IF subunits form coiled-coil dimers
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Heterodimers overlap to form filamentous tetramers

• Coiled-coils align to form antiparallel staggered
  structures

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Assembly of a mature IF from tetramers occurs in
3 stages
Figure 05.05 A model for intermediate filament
assembly. The coiled coil formed by the dimer
formed the structural basis for the strength of
intermediate filaments.
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Posttranslational modifications control the
shape of intermediate filaments

• Chemical modification of IF controls their shape
  and function
• Phosphorylation-dephosphorylation
• Glycosylation
• Farnesylation
• Transglutamination of head and tail domains

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IFs form specialized structures

• Keratins in epithelium

• Costameres

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Microtubules (MT) organize movement inside a cell

• Key Concepts (1)
• MTs are hollow, tube-shaped polymers comprised of
  proteins called tubulins.
• MTs serve as roads or tracks that guide the
  intracellular movement of cellular contents.
• MT formation is initiated at specific sties in
  the cytosol called MT-organizing centers. The
  basic building block of a MT is a dimer of two
  different tubulin proteins.

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MTs organize movement inside a cell

• Key Concepts (2)
• MTs have structural polarity, which determines
  the direction of the molecular transport they
  support. This polarity is caused by the binding
  orientation of the proteins in the tubulin dimer.
• The stability of MTs is determined, at least in
  part, by the type of guanine nucleotides bound by
  the tubulin dimers within it.
• Dynamic instability is caused by the rapid growth
  and shrinkage of MTs at one end, which permits
  cells to rapidly reorganize their MTs.

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MTs organize movement inside a cell

• Key Concepts (3)
• MT-binding proteins play numerous roles in
  controlling the location, stability, and function
  of microtubules.
• Dyneins and kinesins are the motor proteins that
  use ATP energy to transport molecular cargo
  along MTs.
• Cilia and flagella are specialized MT-based
  structures responsible for motility in some
  cells.

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MT cytoskeleton is a network of "roads" for
molecules "pass to and fro"
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MT assembly begins at a MT-organizing center
(MTOC)
Figure 05.08 The distribution of microtubules in
a human epithelial cell. The microtubules are
stained green and the DNA is stained red.
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The MTOC contains the gamma tubulin ring complex
(?TuRC) that nucleates MT formation

• Centrioles
• Pericentriolar material
• gamma (? ) tubulin  

Figure 05.09 The structure and location of the
centrosome.
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The primary building block of MTs is an
alpha-beta tubulin dimer

• a - and ß -tubulin bind together to form stable
  dimer
• If purified a-ß  tubulin dimers bound to GTP are
  concentrated enough (critical concentration),
  they spontaneously form MTs  

Figure 05.10 A three dimensional model of the
dimer formed by a- and ß-tubulin.
Figure 05.11 In vitro assembly of microtubules
is spontaneous and GTP-dependent. The graph
represents the turbidity of a solution of a-ß
tubulin dimers over time.
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MTs are hollow "tubes" composed of 13
protofilaments

• Polymers of dimers ? sheet composed of 13
  protofilaments ? folds into a tube
• GTP binding and hydrolysis regulate MT
  polymerization and disassembly

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The growth and shrinkage of MTs is called dynamic
instability

• Some microtubules rapidly grow and shrink in
  cells dynamic instability
• Elongation is at the
• end by GTP-bound dimers

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Catastrophe?

• What happens when the supply of GTP-bound tubulin
  dimers runs out?
• 1) MT depolymerizes at the end
• OR
• 2) Capping proteins prevent depolymerization

Figure 05.15 Two fates of the plus ends of
microtubules.
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Some MTs exhibit treadmilling

• In cases where neither end of MT is stabilized,
  tubulin dimers are added to the end and lost
  from the - end
• Overall length of these MTs remains fairly
  constant, but the dimers are always in flux

Figure 05.16 Treadmilling in microtubules.
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Benefits of dynamic instability

• Allows cells to have
• flexibility with trafficking during cell movement
• ability to exert force by bonding with cargo
  molecules

Figure 05.17 Microtubules exert enough force to
move cargo by dynamic instability.
Figure 05.18 Longitudinal and lateral bonds make
microtubules strong.
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MT-associated proteins regulate the stability
and function of MTs

• MAPs capping proteins, rescue-associated
  proteins, and proteins that govern the motion
• motor protein special type of MAP that
  transports organelles/vesicles
• Dyneins and kinesins

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Motors
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Cilia and Flagella

• Axoneme

• Sliding dynein whip movement

Figure 05.23 The structure of an axoneme.
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Actin filaments control the movement of cells

• Key Concepts (1)
• Actin filaments are thin polymers of actin
  proteins.
• Actin filaments are responsible for large-scale
  changes in cell shape, including most cell
  movement.
• Actin filament polymerization is initiated at
  numerous sites in the cytosol by actin-nucleating
  proteins.
• Actin filaments have structural polarity, which
  determines the direction that force is exerted on
  them by myosin motor proteins.

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Actin filaments control the movement of cells

• Key Concepts (2)
• The stability of actin filaments is deteremined
  by the type of adenine nucleotides bound by the
  actin proteins within them.
• Actin-binding proteins play numerous roles in
  controlling the location, stability, and function
  of actin filaments.
• Cell migration is a complex process, requiring
  assembly and disassembly of different types of
  actin filament networks.

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The building block of actin filaments is the
actin monomer

• Smallest diameter of cytoskeletal filaments 7nm
  microfilament
• Great tensile strength
• Structural polarity
• end barbed end
• - end pointed end
• Often bound to myosin

Figure 05.25 The general structure of an actin
filament. The lateral and longitudinal bonds
holding actin monomers together are indicated at
right.
Figure 05.26 An electron micrograph of an actin
filament partially coated with mysoin proteins.
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Actin found in wide variety of locations and
configurations
Figure 05.27 A number of different actin
filament-based structures in cells.
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ATP binding/hydrolysis regulate actin filament
polymerization and disassembly

• ATP polymerization
• ATP?ADP depolymerization

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Actin polymerization occurs in 3 stages
Figure 05.29 The three stages of actin filament
assembly in vitro.
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Actin filaments have structural polarity

• Actin filaments undergo treadmilling

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6 classes of proteins bind to actin to control
its polymerization/organization

1. Monomer-binding proteins regulate actin
   polymerization
2. Nucleating proteins regulate actin polymerization

Figure 05.31 The structure and function of
profilin, an actin monomer-binding protein.
Figure 05.32 ARP2/3 nucleates the formation of a
new actin filament off the side of an existing
filament.
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6 classes of proteins bind to actin to control
its polymerization/organization

• 3. Capping proteins affect the length and
  stability of actin filaments
• 45. Severing and depolymerizing proteins control
  actin filament disassembly
• 6. Cross-linking proteins organize actin
  filaments into bundles and networks

Figure 05.34 Three forms of crosslinked actin
filaments created by different crosslinking
proteins.
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Cell Migration

• Actin-binding motor proteins exert force on actin
  filaments to induce cell movement
• Cell migration is a complex, dynamic
  reorganization of an entire cell
• Migrating cells produce three characteristic
  forms of actin filaments filopodia,
  lamellopodia, and contractile filaments

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Filopodia
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Myosins are a family of actin-binding motor
proteins

• myosins multisubunit proteins organized into 3
  structural domains
• Motor
• Regulatory
• Tail

Figure 05.36 Myosin proteins contain three
funtional domains
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Contractile cycle

• Myosins move towards one end of the actin
  filaments
• myosin V crawls towards the - end, all other
  myosins crawl towards the end
• Allows for movement of cell

Figure 05.37 The contractile cycle of myosin.
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Striated muscle contraction is a well-studied
example of cell movement
Figure 05.38 The anatomy of a skeletal muscle.
The sarcomere contains actin and myosin arranged
in parallel bundles.
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Eukkaryotic cytoskeletal proteins arose from
prokaryotic ancestors

• Modern prokaryotic cells express a number of
  cytoskeletal proteins that are homologous to
  eukaryotic cytoskeletal proteins and behave
  similarly
• Vimentin (IF)
• FtsZ (MT)
• MreB and ParM (actin)
• Shared properties seem to include protection of
  DNA, compartmentalization and motility.