Contraction, Locomotion (Ch 16), Shelden
Actin-dependent ATP hydrolysis by myosin generates motile force
Molec Biol Cell 5th ed. Supplemental Videos
1
The Myosin ATPase activity cycle
ATP binding releases myosin from actin (rigor is caused by lack of ATP)
ATP hydrolysis to ADP+Pi moves myosin forward
Release of Pi accompanies reattachment of myosin to a new actin subunit
Release of ADP causes force generation and movement
Fig 16-61
2
The Myosin ATPase activity cycle
Molec Biol Cell 5th ed. Supplemental Videos
3
Different Myosins Exhibit Different Motile Behaviors
All myosins except myosin VI move toward the plus
end of actin filaments
Fig 16-57
4
Contractile force is generated in all types of cells
by the action of bipolar myosin II thick filaments
Fig 16-54
Fig 16-55
5
Smooth muscle cells (and non-muscle cells) assemble myosin thick
filaments as needed by regulating myosin light chains
Fig 16-72
6
Myosin thick filaments assemble during smooth muscle cell
contraction and apply force to preassembled actin filaments
Actin filaments are attached to the cell membrane at dense plaques containing alpha
actinin and other actin associated proteins. Fujiwara et al., J. Cell Biol (1983) 96:783-795
Anti alpha actinin
7
Isolated smooth muscle cell contraction
relaxedformation of myosin
thick filaments contraction
Driska et al., Journal of Applied Physiology, Vol. 86,
Issue 1, 427-435, January 1999
8
Formation of skeletal (striated) muscle
Skeletal muscle myotube
Showing nucleiMyotube formation from myoblasts
9
Skeletal muscle cells contain thick filaments
organized into permanent sarcomeres
Fig 16-74
Fig 16-74
I-band A-band
10
Sarcomeres are highly ordered structures
Actin thin filament length is determined by nebulin
The plus ends of actin are attached to Z discs and
capped by CapZ
Minus ends of actin are protected by tropomodulin
Thick filaments are positioned by titin
Fig 16-75Fig 16-76
11
Thick and thin filaments slide past each other
during contraction, but don’t change length.
Fig 16-76
12
Regulation of sarcomere contraction
Nerve impulses cause release of acetylcholine at the junction. Binding of
acetylchline to the muscle cells opens ion changels that lead to depolarization of the
plasma membrane (sarcolema) and transverse tubules
Neuromuscular junctionsFig 16-77
13
Depolarization of the tranverse tubules stimulates release of calcium from the
sarcoplasmic reticulum (a modified ER) into the cytoplasm
Fig 16-77
14
Calcium binds to the troponin complex and causes
attached tropomyosin to move away from actin
Troponin C: binds calcium
Troponin T: attaches
tropomyosin
Troponin I: inhibits myosin
activity by changing
shape
Myosin binding sites on
actin are revealed by
movement of
tropomyosinFig 16-78
15
Smooth and Striated Muscle
Smooth Striated
Actin meshwork anchored in dense
plaques by alpha-actinin. Myosin
not assembled when relaxed.
Individual cells Multicellular syncitia (skeletal)
Actin bundles anchored in Z lines
using alpha-actinin. Myosin
assembled in thick filaments.
Simple cell architecture Myofibrils with M-lines, Z-lines, A bands,
I bands. Transverse tubules and
sarcoplasmic reticulum
Contraction initiated by activation
of myosin light chain kinase, causes
mini-myosin filaments to form
Neurons depolarize sarcolema.
Depolarization enters T-tubule system,
causing calcium release from sarcoplasmic
reticulum. Calcium binds to Troponin
complex, moving tropomyosin out of the
myosin binding site on actin filaments.
Cells
Actin
Organiz
ation
Trigger
16
Movement of cells involves protrusion, attachment, transport of
the cell components and retraction of trailing attachments.
Fig 16-86
17
Amoeboid cell movement
Molec Biol Cell 5th ed. Supplemental Videos
18
Keratocyte movie
Rachael Ream, George Somero & Julie Theriot.
http://cmgm.stanford.edu/theriot/movies.htm#Current
19
Membrane protrusion is driven by actin
polymerization
Lysteria movement (J. Theroit, Ph.D.)
20
Cell Locomotion
Lysteria movement (J. Theroit, Ph.D.)
http://cmgm.stanford.edu/theriot/IFLmBig.jpg
21
Protrusion
Lamellar (and pseudopod)
protrusion of the plasma
membrane is driven by
cyclical actin polymerization
at the cell periphery and
depolymerization in more
interior sites
Cofilin prefers ADP
actin
22
Attachment is mediated by formation
of “focal adhesions”
Fig 19-45
Elements of Focal Adhesions:
1. Transmembrane proteins
(integrins)
2. Regulatory elements
3. Coupling proteins talin and
vinculin
4. Actin
23
Crawling movement of fibroblasts
Copyright Eric Shelden
24
Focal adhesions in a moving
fibroblast
Bhatt et al., Journal of Cell Science 115, 3415-3425 (2002)
25
Forward transport of organelles is mediated by
motor proteins (myosins, dyneins and kinesins)
Wu et al., Cell Biol., Volume 143, Number 7,
December 28, 1998 1899-1918
See also http://micro.magnet.fsu.edu/moviegallery/pondscum/protozoa/amoeba/index.html
26
Fig 16-94
Retraction of trailing cellular attachments is mediated
by myosin II-dependent contraction at the rear of cells
Myosin I (green) and Myosin
II (red) in a motile cell
Da
vid K
necht, U
nive
rsity o
f Conne
cticut
Wild-t
ype a
moeba
Myosi
n II
nul
ls
27
(Video on next slide)
Video