Actin, Myosin, and Cell Movement

Actin, Myosin, and Cell Movement
 Actin filaments, usually in association with myosin, are
responsible for many types of cell movements.
 Myosin is the prototype of a molecular motor—a protein that
converts chemical energy in the form of ATP to mechanical
energy, thus generating force and movement.
 The most striking variety of such movement is muscle
contraction, which has provided the model for
understanding actin-myosin interactions and the motor activity
of myosin molecules.
 Interactions of actin and myosin are responsible not only for
muscle contraction but also for a variety of movements of
nonmuscle cells, including cell division, so these interactions play
a central role in cell biology.
 Moreover, the actin cytoskeleton is responsible for the crawling
movements of cells across a surface, which appear to be driven
directly by actin polymerization as well as actin-myosin
interactions.
Muscle Contraction
 Muscle cells are highly specialized for a single task, contraction.
 There are three distinct types of muscle cells in vertebrates:
 skeletal muscle, which is responsible for all voluntary
movements;
 cardiac muscle, which pumps blood from the heart; and
 smooth muscle, which is responsible for involuntary movements
of organs such as the stomach, intestine, uterus, and blood
vessels.
 In both skeletal and cardiac muscle, the contractile elements of
the cytoskeleton are present in highly organized arrays that give
rise to characteristic patterns of cross-striations.
It is the characterization of these structures in skeletal muscle that has
led to our current understanding of muscle contraction, and
other actin-based cell movements, at the molecular level.
Actin myosin cell movement.doc; Page 1 of 16

Skeletal muscles are bundles of muscle fibers, which are single large
cells (approximately 50 μm in diameter and up to several centimeters
in length) formed by the fusion of many individual cells during
development. (Figure 11.18)
 Most of the cytoplasm consists of myofibrils, which are cylindrical
bundles of two types of filaments: thick filaments
of myosin (about 15 nm in diameter) and thin filaments
of actin (about 7 nm in diameter).
 Each myofibril is organized as a chain of contractile units
called sarcomeres, which are responsible for the striated
appearance of skeletal and cardiac muscle.
 The sarcomeres (which are approximately 2.3 μm long) consist
of several distinct regions, discernible by electron microscopy,
which provided critical insights into the mechanism of muscle
contraction (Figure 11.19).

 The ends of each sarcomere are defined by the Z disc.
 Within each sarcomere, dark bands (called A bands because they
are anisotropic when viewed with polarized light) alternate with
light bands (called I bands for isotropic).
 These bands correspond to the presence or absence
of myosin filaments.
 The I bands contain only thin (actin) filaments, whereas the A
bands contain thick (myosin) filaments.
The myosin and actin filaments overlap in peripheral regions of the A
band, whereas a middle region (called the H zone) contains only
myosin.
 The actin filaments are attached at their plus ends to the Z disc,
which includes the crosslinking protein α-actinin.
 The myosin filaments are anchored at the M line in the middle of
the sarcomere.
Two additional proteins (titin and nebulin) also contribute to sarcomere
structure and stability (Figure 11.20).
 Titin is an extremely large protein (3000 kd), and single titin
molecules extend from the M line to the Z disc.
 These long molecules of titin are thought to act like springs that
keep the myosinfilaments centered in the sarcomere and
maintain the resting tension that allows a muscle to snap back if
overextended.
 Nebulin filaments are associated with actin and are thought to
regulate the assembly of actin filaments by acting as rulers that
determine their length.

Sliding Filament Model:
The basis for understanding muscle contraction is the sliding filament
model, first proposed in 1954 both by Andrew Huxley and Ralph
Niedergerke and by Hugh Huxley and Jean Hanson (Figure 11.21).
 During muscle contraction, each sarcomere shortens, bringing
the Z discs closer together.
 There is no change in the width of the A band, but both the I
bands and the H zone almost completely disappear.
 These changes are explained by the actin andmyosin filaments
sliding past one another, so that the actin filaments move into
the A band and H zone.
 Muscle contraction thus results from an interaction between
the actin and myosin filaments that generates their movement
relative to one another.
The molecular basis for this interaction is the binding of
myosin to actin filaments, allowing myosin to function as a motor that
drives filament sliding.
The type of myosin present in muscle (myosin II) is a very large
protein (about 500 kd) consisting of two identical heavy chains (about
200 kd each) and two pairs of light chains (about 20 kd each) (Figure
11.22).

 Each heavy chain consists of a globular head region and a
long α-helical tail.
 The α-helical tails of two heavy chains twist around each other in
a coiled-coil structure to form a dimer, and two light chains
associate with the neck of each head region to form the
complete myosin II molecule.
 The thick filaments of muscle consist of several
hundred myosin molecules, associated in a parallel staggered
array by interactions between their tails (Figure 11.23).
 The globular heads of myosin bind actin, forming cross-bridges
between the thick and thin filaments.
 It is important to note that the orientation of myosin molecules
in the thick filaments reverses at the M line of the sarcomere.
 The polarity of actin filaments (which are attached to Z discs at
their plus ends) similarly reverses at the M line, so the relative
orientation of myosin and actin filaments is the same on both
halves of the sarcomere.
 The motor activity of myosin moves its head groups along
the actin filament in the direction of the plus end.
 This movement slides the actin filaments from both sides of the
sarcomere toward the M line, shortening the sarcomere and
resulting in muscle contraction.
 In addition to binding actin, the myosin heads bind and
hydrolyze ATP, which provides the energy to drive filament
sliding.

 This translation of chemical energy to movement is mediated by
changes in the shape of myosin resulting from ATP binding.
The generally accepted model (the swinging-cross-bridge model) is
that ATP hydrolysis drives repeated cycles of interaction
between myosin heads and actin.
 During each cycle, conformational changes in myosin result in
the movement of myosin heads along actin filaments.
 Although the molecular mechanisms are still not fully
understood, a plausible working model for myosin function has
been derived both from in vitro studies of myosin movement
along actin filaments (a system developed by James Spudich and
Michael Sheetz) and from determination of the three-dimensional
structure of myosin by Ivan Rayment and his
colleagues (Figure 11.24).

 The cycle starts with myosin (in the absence of ATP) tightly
bound to actin.

 ATP binding dissociates the myosin-actin complex and the
hydrolysis of ATP then induces a conformational change
in myosin.
 This change affects the neck region of myosin that binds the
light chains (see Figure 11.22), which acts as a lever arm to
displace the myosin head by about 5 nm.
 The products of hydrolysis (ADP and Pi) remain bound to
the myosin head, which is said to be in the “cocked” position.
 The myosin head then rebinds at a new position on
the actin filament, resulting in the release of ADP and Pi and
triggering the “power stroke,” in which the myosin head returns
to its initial conformation, thereby sliding the actin filament
toward the M line of the sarcomere.
Regulation of muscle contraction in striated
muscles
The contraction of skeletal muscle is triggered by nerve impulses,
which stimulate the release of Ca2+ from the sarcoplasmic reticulum—
a specialized network of internal membranes, similar to the
endoplasmic reticulum, that stores high concentrations of Ca2+ ions.
 The release of Ca2+ from the sarcoplasmic reticulum increases
the concentration of Ca2+ in the cytosol from approximately 10-
7 to 10-5M.
 The increased Ca2+ concentration signals muscle contraction via
the action of two accessory proteins bound to
the actin filaments: tropomyosin and troponin (Figure 11.25).

 Tropomyosin is a fibrous protein that binds lengthwise along the
groove of actin filaments.
 In striated muscle, each tropomyosin molecule is bound to
troponin, which is a complex of three polypeptides: troponin C
(Ca2+-binding), troponin I (inhibitory), and troponin T
(tropomyosin-binding).
 When the concentration of Ca2+ is low, the complex of the
troponins with tropomyosin blocks the interaction
of actin and myosin, so the muscle does not contract.
 At high concentrations, Ca2+ binding to troponin C shifts the
position of the complex, relieving this inhibition and allowing
contraction to proceed.
Contractile Assemblies of Actin and Myosin in
Nonmuscle Cells
Contractile assemblies of actin and myosin, resembling small-scale
versions of muscle fibers, are present also in nonmuscle cells.

 As in muscle, the actin filaments in these contractile assemblies
are interdigitated with bipolar filaments of myosin II, consisting
of 15 to 20 myosin II molecules, which produce contraction by
sliding the actin filaments relative to one another (Figure 11.26).
 The actin filaments in contractile bundles in nonmuscle cells are
also associated with tropomyosin, which facilitates their
interaction with myosin II, probably by competing with filamin
for binding sites on actin.
 Two examples of contractile assemblies in nonmuscle cells,
stress fibers and adhesion belts, were discussed earlier with
respect to attachment of the actin cytoskeleton to regions of
cell-substrate and cell-cell contacts (see Figures
11.13 and 11.14).
 The contraction of stress fibers produces tension across the cell,
allowing the cell to pull on a substrate (e.g., the extracellular
matrix) to which it is anchored.
 The contraction of adhesion belts alters the shape of epithelial
cell sheets: a process that is particularly important during
embryonic development, when sheets of epithelial cells fold into
structures such as tubes.
 The most dramatic example of actin-myosin contraction in
nonmuscle cells, however, is provided by cytokinesis—the
division of a cell into two following mitosis (Figure 11.27).

 Toward the end of mitosis in animal cells, a contractile
ring consisting of actin filaments and myosin II assembles just
underneath the plasma membrane.
 Its contraction pulls the plasma membrane progressively inward,
constricting the center of the cell and pinching it in two.
 Interestingly, the thickness of the contractile ring remains
constant as it contracts, implying that actin filaments
disassemble as contraction proceeds.
 The ring then disperses completely following cell division.
Regulation of contraction in non-muscle and
smooth muscles
The regulation of actin-myosin contraction in striated muscle,
discussed earlier, is mediated by the binding of Ca2+ to troponin.
 In nonmuscle cells and in smooth muscle, however, contraction
is regulated primarily by phosphorylation of one of
the myosin light chains, called the regulatory light chain (Figure
11.28).

 Phosphorylation of the regulatory light chain in these cells has at
least two effects:
 It promotes the assembly of myosin into filaments, and it
increases myosin catalytic activity, enabling contraction to
proceed.
 The enzyme that catalyzes this phosphorylation,
called myosin light-chain kinase, is itself regulated by association
with the Ca2+-binding protein calmodulin.
 Increases in cytosolic Ca2+ promote the binding of calmodulin to
the kinase, resulting in phosphorylation of the myosin regulatory
light chain.
 Increases in cytosolic Ca2+ are thus responsible, albeit
indirectly, for activating myosin in smooth muscle and
nonmuscle cells, as well as in striated muscle.
Unconventional Myosins
In addition to myosin II (“conventional” two-headed myosin), several
other types of myosin are found in nonmuscle cells.
 In contrast to myosin II, these “unconventional” myosins do not
form filaments and therefore are not involved in contraction.
 They may, however, be involved in a variety of other kinds of
cell movements, such as the transport of membrane vesicles and
organelles along actin filaments, phagocytosis, and extension of
pseudopods in amoebae (see Figure 11.17).

 The best-studied of these unconventional myosins are members
of the myosin I family (Figure 11.29).
 The myosin I proteins contain a globular head group that acts as
a molecular motor, like that of myosin II.
 However, members of the myosin I family are much smaller
molecules (about 110 kd in mammalian cells) that lack the long
tail of myosin II and do not form dimmers.
 Their tails can instead bind to other structures, such as
membrane vesicles or organelles.
 The movement of myosin I along an actin filament can then
transport its attached cargo.
 One function of myosin I, discussed earlier, is to form the lateral
arms that link actin bundles to the plasma membrane of
intestinal microvilli (see Figure 11.16).
 In these structures, the motor activity of myosin I may move the
plasma membrane along the actin bundles, toward the tip of the
microvillus.
 Additional functions of myosin I may be in the transport of
vesicles and organelles along actin filaments and in movement of
the plasma membrane during phagocytosis and pseudopod
extension.
 In addition to myosins I and II, at least 12 other classes of
unconventional myosins (III through XIV) have been identified.
 Some of these unconventional myosins are two-headed
like myosin II, whereas others are one-headed like myosin I.
 The functions of most of these unconventional myosins remain to
be determined, but some have been clearly shown to play
important roles in organelle movement (myosins V and VI) and

in sensory functions such as vision (myosin III) and hearing
(myosins VI and VII).
Cell Crawling
The crawling movements of cells across a surface represent a basic
form of cell locomotion, employed by a wide variety of different kinds
of cells.
 Examples include the movements of amoebas, the migration of
embryonic cells during development, the invasion of tissues by
white blood cells to fight infection, the migration of cells involved
in wound healing, and the spread of cancer cells during the
metastasis of malignant tumors.
 Similar types of movement are also responsible for phagocytosis
and for the extension of nerve cell processes during development
of the nervous system.
 All of these movements are based on the dynamic properties of
the actin cytoskeleton, although the detailed mechanisms
involved remain to be fully understood.
Cell crawling involves a coordinated cycle of movements, which can be
viewed in three stages.
 First, protrusions such as pseudopodia, lamellipodia, or
microspikes (see Figure 11.17) must be extended from the
leading edge of the cell (Figure 11.30).

 Second, these extensions must attach to the substratum across
which the cell is migrating. Finally, the trailing edge of the cell
must dissociate from the substratum and retract into the cell
body.
 A variety of experiments indicate that extension of the leading
edge involves the polymerization and crosslinking
of actin filaments.
 For example, inhibition of actin polymerization (e.g., by
treatment with cytochalasin) blocks the formation of cell surface
protrusions.
 The regulated turnover of actin filaments, as illustrated in Figure
11.5, leads to the extension of processes such as filopodia and
lamellipodia at the leading edge of the cell, and both cofilin and
Arp2/3 proteins appear to be involved in this process.
 Unconventional myosins may also participate in the extension of
processes at the leading edge:
 Myosin I is required for pseudopod extension in the
amoeba Dictyostelium and Myosin V for extension of filopodia in
neurons.
 Following their extension, protrusions from the leading edge
must attach to the substratum in order to function in cell
locomotion.

 For slow-moving cells, such as fibroblasts, attachment involves
the formation of focal adhesions (see Figure 11.13).
 Cells moving more rapidly, such as amoebas or white blood cells,
form more diffuse contacts with the substratum, the molecular
composition of which is not known.
 The third stage of cell crawling, retraction of the trailing edge, is
the least understood.
 The attachments of the trailing edge to the substratum are
broken, and the rear of the cell recoils into the cell body.
 The process appears to require the development of tension
between the front and rear of the cell, generating contractile
force that eventually pulls the rear of the cell forward.
 This aspect of cell locomotion is impaired in mutants
of Dictyostelium lacking myosin II, consistent with a role
for myosin II in contracting the actin cortex and generating the
force required for retraction of the trailing edge.

Actin, Myosin, and Cell Movement

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Actin, Myosin, and Cell Movement