Chapter 9 Cytoskeleton
Microtubule assembly and dynamic instability
GTP required for microtubule assembly. Microtubule disassembly is a rapid process that does not require an input of energy (this spontaneous disassembly is referred to as dynamic instability) DI occurs when incorporation of new-GTP bound dimers is slower than the rate of GTP hydrolysis
Cytoplasmic dynein
Huge protein with a globular,force-generating (ATP hydrolyzing) head. Stalks that interact with microtubule. Stems connected to intermediate and light chains that bind necessary accessory proteins to interact with membrane-bounded cargo
3 kinds of molecular motors
Kinesin and Dynein- move along microtubule tracks. Myosin-moves along microfilament tracks
Major functions of cytoskeleton
Scaffold providing structural support and maintaining cell shape. Internal framework to organize and position organelles. Directs cellular locomotion and movement of materials within a cell. Dynamic change in cell shape and principle force of cell motility
Sliding filament model of muscle contraction
Skeletal muscles contract by myosin motors sliding antiparallel actin filaments; oppositely oriented heads of myosin pull toward the opposing plus ends; A bands remain constant in length; H and I bands decrease in width as Z discs are pulled toward sarcomere center. Muscles do not contract until neuronal impulse is received, releasing calcium from sarcoplasmic reticulum stores
WASP proteins
activated by stimulus at cell surface; recruit and activate Arp2/3 protein complex; initiates nucleation and polymerization of actin; diverse actin-binding proteins, such as profilin, are also activated
Z line
anchor proteins that form the boundaries b/n sarcomeres; where actin filament plus ends are bound, minus ends extend toward sarcomere center
Centrosome
animal specific structures that initiate microtubule nucleation and regulate their dynamics. Each contains two barrel shaped centrioles which are surrounded by pericentriolar material (PCM)
Tropomyosin
associated with resting state actin filaments; occupies gap b/n two actin strands and inhibits myosin from biding to actin filaments. In unstimulated state, it blocks the myosin binding sites on actin monomers
treadmilling
at low ATP concentrations, filaments reach a steady state where addition to the plus end equals loss from minus end
Actin filament depolymerizing proteins
bind actin-ADP subunits at the minus end promoting rapid turnover of actin filaments
Axoneme
central microtubule core found in both cilia and flagella; consists of microtubules in a 9+2 arrangement with diverse accessory proteins. They are nucleated by basal bodies (that produce gammaTURK complexes) and plus end extends into membrane surrounded cilia or flagella
basal bodies
centrioles found at the base of eukaryotic cilia and flagella
Neuromuscular junction
contact between nerve and muscle
plectin
cross bridging protein, connects IFs to other components of cytoskeleton, organelles, and nucleus
Microtubule-Associated Proteins (MAPs)
diverse group of proteins that attach to microtubules an function to increase or decrease stability by promoting either polymerization or disassembly
filopodia
dynamic probing extensions whose membrane tips possess receptor proteins that bind chemoattractants and repellants; this stimulation leads to regulatory changes in cytoskeleton and direction of axonal growth
protofilaments
each microtubule contains an arrangement of 13 of these aligned longitudinally along its length. It is assembled from heterodimers of non-identical alpha and beta-tubulin subunits
Cilia and flagella
hair-like motile organelles that project from PM of diverse eukaryotic cells; distinguished by the differences in their beat processes. Cilia tend to occur in large #s on cell surface, while flagellated cells usually contain only one flagellum. Cilia promote motility in many single-celled organisms, but in multicellular animals they function to circulate extracellular fluids
intermediate filaments
heterogeneous group of proteins that assemble as strong, flexible, rope-like fibers that provide mechanical strength; far less dynamic than microtubules and microfilaments, and much less sensitive to chemical agents that disrupt their structure-highly stable. Five major classes, 1-4 are cytoplasmic while type 5 (lamins) are present in the inner lining of nuclear envelope. Assembly of IFs is spontaneous does not require ATP or GTP hydrolysis . Formed by two antiparallel dimers which causes IFs and tetramers to lack polarity-cannot be used as tracks for motor proteins
Axonemal dynein
hydrolyzes ATP, providing the mechanical forces that promote both cilia and flagellar movement
cortex
in non-muscle motile cells, majority of actin is restricted to this thin region just beneath the PM; here dynamic changes in actin polymerization facilitate cell shape changes that promote crawling and phagocytosis; diverse actin-binding proteins affect localized assembly/disassembly of actin filaments for this purpose
Intraflagellar transport
kinesin and dynein transport materials into and out of both growing and mature structures promoting growth and maintenance, as well as cilia mediate cell-cell signaling
lamellipodium
leading edge along the substrate
Membrane-binding proteins
link microfilaments to proteins in the to PM
gamma-tubulin
localized to PCM and is critical for MT nucleation. Forms ring structures in the PCM known as gamma-TURKs (gamma-tubulin ring complexes that are same diameter as a microtubule. Serves as protective cap to the minus end, preventing its disassembly. gamma-TURKs can be assembled on established microtubules in order to promote branching
M line
middle of the sarcomere where tails of antiparallel myosin fibers overlap
myosins
molecular motors of actin filaments; share a characteristic motor head for binding actin and hydrolyzing ATP; tail is divergent, allowing binding to diverse structures/molecules; approx. 40 are encoded by human genome; two groups
processive movement
motor protein moves in a hand-over hand manner along an individual microtubule for a long distance w/o falling off
Axonal transport
movement of neurotransmitters along length of cell, movement away from cell body and toward cell body, establish tracks for a variety of unidirectional motor proteins
Sarcomere
muscle contractile units with a stereotypical arrangement of actin and myosin fibers; each has a distinct banding pattern due to thick filaments (myosin) and thin filaments (actin)
Cytoskeleton
network of filamentous protein structures: microtubules, microfilaments, and intermediate filaments
Unconventional myosins
often have single head and do not assemble into filaments, but dimers; function with microtubule motors to deliver targets to correct locations; similar to kinesin, unconventional myosin involved in vesicle transport moves in hand over hand fashion
traverse (T) tubules
plasma membrane involutions that connect to the SR; action potential in muscles is propagated into cell interior by this
Cross-linked proteins
possess 2 or more actin binding sites and join microfilaments into 3D networks
Conventional type II myosins
principle motor proteins that promote muscle contraction; all migrate toward barbed plus end pf microfilaments; non-muscle activities include cell division, tensile strength of focal adhesions, cell migration, and direction of axonal growth cone movement. All motor activity is confined to head domain. Each composed of 4 regulatory light chains and two heavy chains associated with catalytic heads
Monomer-polymerizing proteins
promote the growth of actin filaments; the polymerizing protein profiling binds to actin-ADP-monomers and facilitates replacement with ATP, enhancing polymerization
titin
protein and is a giant elastic fiber that extends from the Z line to the M band providing elasticity, resting tension and preventing sarcomere form being pulled apart during muscle stretching
+TIPS
regulated expression and targeting of proteins; can stabilize the plus ends of microtubules and inhibit catastrophe either directly or by facilitating the connection of microtubules to various cellular structures
Dynein
retrograde minus end-directed microtubule movement of vesicles and organelles. Responsible for movement of cilia and flagella
rhodamine
small, fluorescent dye that can be covalently linked to proteins
Microtubule-Organizing Centers( MTOCs)
specialized structures for nucleation and assembly of microtubules
minus end
terminates with alpha-subunits and is static-no growth or disassembly; pointed and has lower affinity for monomers
A band
thick filaments; myosin filaments engaging with actin filaments
growth cones
tip of migrating neuronal axons; within these lamellipodia reside probing extensions known as filopodia (microspikes)
Sliding-microtubule mechanism
tubules are fixed in place by radial spokes and the basal body; when dynein hydrolyzes ATP and attempts to "walk" toward the minus ends of associated B tubules, this causes adjacent microtubules to slide past one another. Through precise controlling of when and which dyneins hydrolyze ATP, distinct patterns of ciliary and flagellar movements are coordinated
Two groups of myosin
1.) Conventional (type II) 2.) Unconventional myosins (types 1, 3-18)
motor proteins
ATP-hydrolysis protein machine that bind specific cargo, like vesicles or organelles, via adapter proteins
Troponin
associated with resting state actin filaments; binds and links actin and tropomyosin; following impulse it binds to released Ca2+, causing a conformational change that shifts tropomyosin exposing myosin binding sites
how is microfilament cytoskeleton organized?
by controlling equilibrium between assembly and disassembly of microfilaments
Filament-severing proteins
cleave filaments internally
Myofibril
composes muscle fibers; consists of repeating array or sarcomeres
plus-end
end of microtubule terminates with beta-subunits. This is the dynamic end where growth and disassembly occurs; higher affinity for monomers and incorporates them about 10x as fast as other end
Focal adhesions
formed via localized integrals, providing anchorage support
microtubules
hollow, cylindrical rod-like structures
Keratin IFs
major structural proteins of epithelial cells establishing a protective barrier in such tissues
neurofilaments
major supporting network in neuronal axons
Kinesin
member of a superfamily called Kinesin-related proteins. Each is heterotetramer of two identical heavy and light chains. It is a plus end-directed micro tubular motor (only moves toward plus end of microtubules (away from nucleus))
muscle fibers
multinucleate cells resulting from embryonic fusion of myoblasts;
H zone
myosin tails only (center or sarcomere)
situs inversus
syndorme in which the left-right body symmetry is reversed; often results from mutations in genes encoding ciliary proteins
in vivo
taking place in a live organism
I band
thin filament; actin filaments only
Monomer-sequestering proteins
(negative regulation) bind to actin-ATP monomers and prevent them from polymerizing
End-blocking (capping) proteins
(negative regulation), but also stabilizing. Regulate the length of actin filaments
Nucleating proteins
(positive regulation) microfilament assembly occurs rapidly if filament "seed" is present; these proteins bind monomers and catalyze their polymerization into small filaments through regulation of the Arp2/3 protein complex
Actinomyosin contractile cycle
ATP binds myosin head, conformational change causes detachment of the head from the actin filament. Hydrolysis of ATP changes conformation increasing affinity for actin monomers • Release of Pi further enhances affinity and promotes conforma6onal change on myosin that produces the "power stroke" that moves the thin filament toward the center of the sarcomere. • Release of the ADP allows binding of ATP and repeating the cycle. • In the absence of ATP, myosin remains tightly bound - rigor mortis