Topic 5: The Cytoskeleton Flashcards
What is the structure and function of the cytoskeleton?
movement: subcellular movement, organismal movement (muscles)
organization: polarity, nuclear organization
shape: organellar shape, cellular shape
What are the three filament types in the cytoskeleton?
microfilaments: muscle, cell polarization, cytokinesis
intermediate filaments: nuclear lamino, desmosomes
microtubules: vesicle movement, chromosomal movement
What is the structure of microfilaments?
organize into many subcellular structures: microvilli, cell cortex, adherens belts, stress fibers, contractile ring
composed of actin monomers: many different isoforms that are expressed differentially depending on cell type
G actin: globular actin, monomer
F actin: filamentous, polymer
F actin has structural and functional directionality (positive favors polymerization, negative favors depolymerization)
What are the kinetics of polymerization?
because of structural polarity of the actin filament, the minus end ATP-cleft is exposed to solution and can hydrolyze into ADP, this hydrolysis causes destabilization and depolymerization of the actin filament
How is the actin filament both polymerized and depolymerized?
the actin filament is both polymerizing at the plus end and depolymerizing at the minus end at the same time
G actin can bind ATP or ADP
ATP bound is polymerization favoured
ADP bound is depolymerization favoured
What is actin polymerization in vitro?
ATP-G actin is added to the plus end: if in excess –> polymerization
requires nucleation of polymerization, short oligomers of G-actin are needed to nucleate filament formation
filament formation is must faster at the positive end because of the shielding of the ATP cleft
polymerization is possible at both positive and negative ends with excess ATP-G actin but at steady state occurs at positive end
steady state = treadmilling: balance of pol at positive end and depol at negative is sufficient to generate force
What is actin-associated protein modulate treadmilling?
e.g. actin sequestering proteins: bind available G actin to prevent polymerization
e.g. profilin: enhances the loss of ADP off ADP-G actin, promotes polymerization
What are the motors of microfilaments?
actin dynamics can cause cellular motion but most actin-based movement is caused by myosin motor movement
What is myosin II?
myosins are ATP dependent proteins that can bind actin, undergo a conformational change and move against the rigid actin CSK
all myosins contain: actin binding head, heavy chain (motor unit), flexible neck region, cargo binding tail
type II myosins are hexamers
typical type II myosin consists of: 2 heavy chains that associate by dimerization of tails, 2 essential light chains, 2 regulatory light chains
What is the contractile mechanism?
head domains can bind actin and ATP
neck region can bend in response to the chnaging conformation of the head domain
head domain changes conformation in response to ATP hydrolysis
mechanism of myosin II movement
What is the cross bridge cycle?
release of Pi allows myosin to bind actin
release of ADP that drives “power stroke” = contraction
if not ATP or ADP is bound, the myosin is in a bound and contracted state
myosin is not bound to actin when myosin bonds to ATP, ATP associates releases myosin off actin
ATP hydrolysis allows myosin cocking (opening of myosin)
What are contractile ring in cytokinesis?
non-muscle type II myosin-based constriction during cytokinesis
actin is bundled into a circumferential array at the cortex
pure-string model of contraction
What are microtubules?
structure MT’s are hollow polymers of alpha and beta tubulin dimers
each tubulin subunit also binds GTP and beta tubulin has a GTPase function
the protofilaments have intrinsic polarity (positive and negative ends)
composed of polymers of tubulin dimers
dimers polymerize into protofilaments
tubulin monomers can be GTP bound (pol favoured) or GDP bound (depol favoured)
13 protofilaments assemble into a hollow tube
What is a microtubule organizing centers?
microtubules are anchored in structures called MTOCs in the cell with the negative end anchored into the MTOC (centrosome) which is usually located centrally in the cell close to the nucleus
negative ends are centered near middle of cell
MTOC sits near the interphase nucleus
positive radiate to periphery of cell
negative end of MT are anchored in centrosomes/MTOC
centrosomes contain centrioles and pericentriolar material
What are the kinetics of microtubules?
MTs are continuously assembled and disassembling at the ends and can facilitate CSK-based movements
in vitro: polymerization can occur at both ends (negative end not anchored in MTOC in vitro)
balance of pol/depol is sufficient to generate force
What is treadmilling?
treadmilling in MTs has a greater cellular role than with MFs for cellular movements
less common in MT because negative end is stabilized by centrosome/MTOC
treadmilling generates forces for chromosomal movement in mitosis
What is dynamic instability?
refers to the difference in growing and catastrophe phases in the normal growth of MTs, it is not as simple as just treadmilling
growth of microtubules can occur rapidly at the positive end
growth of MT is fast at positive end as long as concentration of GTP-tubulin is high
polymerized GTP-tubulin forms straight protofilaments
polymerized GDP-tubulin forms curled protofilaments
tubulin has GTPase activity in beta subunit
dynamic instability occurs because of the GTP hydrolysis that occurs in beta tubulin
What are the steps of dynamic instability?
addition of GTP tubulin forma a “GTP cap”
if concentration of GTP tubulin increases, then polymerization at the positive end
if keep adding GTP-tubulin then MT is stabilized because GTP tubulin forms straight protofilaments
if concentration of GTP tubulin decreases then cap is not present and protofilaments curl away = rapid depolymerization = catastrophe
after significant catastrophe, MT can recover, if pool GTP-tubulin is replenished
What are the two types of motors associated with microtubules?
kinesin: positive end directed MT motor
dynein: negative end directed MT motor
What is kinesin?
dimers of heavy chains with association to light chains
monkey-bar movement
What is dyenein?
6 subunits (2 large, 2 intermediate, 2 small)
negative end directed
inch worm movement
How do kinesin and dynein cooperate?
facilitate vesicular transport
coordination of anterograde and retrograde transport
melanophores in fish
allows distribution of pigmentation that is neuronally controlled
camouflage or mating displays
What is the structure of intermediate filaments?
biochemically heterogenous, no true monomer
no associated motor proteins
not involved in cell movement
provide structural integrity to the cell
no intrinsic polarity
What is the function of intermediate filaments in the nuclear lamina?
network of lamin intermediate filaments that forms a meshwork of extending on the inner surface of the nuclear envelope
composed of lamin A, B, and C
provides structural support to the nucleus
sits on the inner face of nuclear envelope
must be disassembled upon mitotic entry (nuclear envelope breakdown)
NEBD is regulated by the phosphorylation of lamins
