Module 5 Flashcards
what is the cytoskseleton
network of structural proteins found in all cell types, defines cell shape and distribution of cellular content
occupies a large portion of the cytosol
permits signalling, vesicular transport and can allow cell motility
what are the classes of structural proteins
- intermediate filaments
- microtubules
- actin
what are intermediate filaments
supply mechanical strength to cells allowing them to resist changes of shape (strongest filament)
are polymers and their expression is tissue and cell specific
primary structure of intermediate filaments
polymer of amino acids link together by peptide bonds
at this stage filaments have the same strength as other proteins in the body
secondary structure of intermediate filaments
rich in alpha helices
- responsible for long, coiled structure of filaments
- hydrogen bonds stabilize structure (resist stretching and prevent collapse)
tertiary and quaternary structure of intermediate filaments
tertiary - coiled monomer
two coiled monomers come together to form a dimer
- monomers wrap around each other forming coiled coil (allows max hydrogen bonding between peptides) giving great strength
two dimers assemble in antiparallel staggered manner forming tetramer
- increase hydrogen bonding and strength
tetramer building block of filament
assembly of intermediate filaments
8 tetramers come together to form a unit length filament (20nm)
unit length filaments come together to form a immature filament (interact loosely end to end)
immature filaments compact to form a mature filament (10nm)
intermediate filaments post translational modifications
help control shape and function
modifcations occur in the head and tail domains of the filament subunit proteins
phosphorylation leads to dissolution of the filament into unit length filaments, when phosphates removed by phosphatases filaments reform (process important for cell division)
what is lamin (intermediate filament)
found solely in nucleus
forms nuclear matrix
dense network to protect chromatin
what is desmin (intermediate filament)
does not form long thin filamentous structure
connects different cellular structures together
important for muscle structure integrity
what is keratin (intermediate filament)
binds to desmosomes to form a complex
makes up hair skin and nails
purpose of microtubules
cellular trafficking
movement of proteins, vesicles and some cellular organelles
create specific routes for cargo, can be assembled/dissembles to create or remove routes
travel can be bi direction and cargo can attach or detach anywhere along length
where does microtubules assembly occur
does not occur spontaneously
assembly required many proteins
occurs in regions called microtubule organizing centres (MTOCs)
assembled in different locations
example of an MTCO is the centrosome (used in cell division)
protein structure of microtubules
made up of tubulins (protein)
alpha and beta tubuline both globular protein with similar shapes that can bind tightly together (head to tail) to form a dimer
both tubulin proteins bind to a GTP molecules
beta tubulin can cleave its GTP to GDP, when bound to GDP beta has a shape change
microtubule polymerization/formation of the tubes
dimers spontaneously assemble into unstable polymers that can quickly fall apart
polymer of 6+ dimers is stable, may grow laterally or longitudinally (protofilament)
protofilaments form sheet and will assemble into a tube of 13 protofilaments
nucleation site for microtubule elongation
at the end of microtubule dimers come and go
rate of assemble greater grows, disassemble greater shortens
microtubule assembly
alpha tubulin always has GTP
beta may have GTP or GDP
when GTP bound to beta, dimer polymerization is favoured and dimers attach to each other
microtubule disassembly
when beta tubulins GTP is hydrolysed to GDP, dimer undergoes conformational change that promotes depolymerization
polarity of microtubule
ends are different, one plus one minus so polar
preference for dimer binding is at plus end so rate is faster there
microtubule dynamic instability
ability to rapidly grow or shrink which is necessary for responses to changes in cellular environment
growing microtubule has a cap of GTP subunits as tip
GTP hydrolysis occasionally exposes GDP bound subunits at tip
rapid catastrophic depolymerization occurs
GTP subunits bind to recap microtubule and stop depolymerization
growth reoccurs when GTP bound dimers available until another change in environment is detected
microtubule catastrophe measures against
when there is rapid depolymerization resulting in shortening
capping
plus ends capping proteins bind adding stability, keep them polymerized even if GDP bound form
rescue
halted or revered, occurs spontaneously is enough GTP dimers present. can occurs in the presence of some other proteins
functions of microtubule based motor proteins
control trafficking
bind to cargo thats needs to be moved then binds to microtubule and walks along it
process consumes ATP
types of microtubule motor proteins
kinesin
moves towards plus end
dynein
moves towards minus end
heads contain microtubule binding domains, have two heads
tails bind to cargo
walking of motor proteins process
head 1 bound to microtubule, head 2 bound to ADP
walking initiated by ATP binding to head 1, conformational change > head 2 swings around
head 2 goes overing binding site on microtubule and binds releasing ADP
head 1 undergoes hydrolysis so ADP bound, release from microtubule
process repeats
actin and microtubules similarities
composition
- globular proteins
movement
- motor proteins used to initiate movement along both proteins
actin and microtubule differences
network formation
- microtubules form dynamic network
- actin forms stronger network that contributes to both structure of the cell and large scale movements (muscle contraction)
actin cytoskeleton can move the cell itself
basic building block of actin
actin monomers
cells can express several different types of actin monomers which allow the cells to match the monomers to their specific functional needs
formation of actin filmaments
actin monomers come together to form long thin actin filaments
bind longitudinally and laterally (high tensile strength can withstand pulling forces that would pull microtubules apart)
actin filaments polarization
there is a plus end (barbed end), and minus end (pointed end)
actin polymerization (what it binds to)
actin monomers bind to nucleotide phosphates (ATP/ADP)
binding of ATP promotes assembly
ADP disassembly
preference for ATP so when there is a constant source ADP is replaced
stages of actin polymerization/formation of filament
nucleation
- two actin monomers dimerize
- nucleation occurs when a third actin monomer binds to the dimer to form a nucleus trimer
- trimer forms core
elongation
- actin monomers added to core, elongates in both directions (plus end favoured)
- dynamic process
steady state
- rate of assembly equals disassembly so net elongation ceases
actin treadmilling
treadmilling is the favoured addition of monomers to one end and the same rate of removal on the other end
keeps filament same length but moves is within the cell
allows cell to rapidly adjust actin cytoskeleton much faster than intermediate filaments which require phosphorylation
when does actin treadmilling occur
regulated by ATP actin concentration compared to ADP bound actin
ATP lower at plus end than minus end
if ATP concentration increases above critical concentration for the minus end, monomers can be added again
acting binding proteins types
- monomer binding proteins
- nucleating proteins
- capping proteins
- severing and depolymerization proteins
- cross linking proteins
- membrane anchors
- actin binding motor proteins
monomer binding proteins
directly bind to actin monomers to influence polymerization
nucleating proteins
bind to actin polymers to increase stability and allow growth of a new branch
capping protein
bind to plus or minus end to stabilize polymer and prevent assembly/disassembly
severing and depolymerization proteins
bind to actin polymer and sever or induce disassembly
cross linking protein
allow side to side linkage of actin polymers to form bundles of actin filaments
membrane anchors
link actin filaments to non actin structural proteins (intergrins)
actin binding motor proteins
bind to actin filament and allow movement
myosin (has 18 different families, each perform specific roles)
myosin structure
motor domain, formed by heavy chain, binds to actin filament and ATP
regulatory domain formed by heavy chain, moves back and forth as myosin moves
tail domain binds to other cellular proteins or myosin
movement of myosin
hydrolysis
- ATP bound to motor domain, myosin is unbound to filament
- hydrolysis of ATP to ADP and inorganic phosphate cause conformational shift in regulatory domain (swing like a lever)
actin binds
- motor domain binds to actin filament
- inorganic phosphate released
- conformational change and pulling myosin along filament
- ADP released, ATP rebind cause myosin to unbind from actin
movement
- myosin moved towards plus end (barbed)
cellular migration
physical movement of a cell
needs lots of coordination to ensure contents stay intact and functional
dynamic assembly/disassembly of cytoskeleton filaments important to generate forces and coordination for migration
process of cellular migration
initiated when actin filaments polymerize near the plasma membrane and push it outwards
pushing forces dont rip the membrane due to hydrophobic interactions between membrane phospholipids
as filopodia and lamellipodia extend plasma membrane bound integrins bind to extracellular matrix
filaments bind to integrins as anchors
types of migration actin filaments
- filopodia
- lamellipodia
- stress fibers
filopodia
thin parallel bundles of filaments
plus end facing membrane
extends in the direction of movement
lamellipodia
larger sheet like bundles of actin filaments
plus end towards membrane
from broader structure that distend a wider amount of plasma membrane in the same direction as filopodia
stress fibers
form around integrins
resemble flipodia but polarity different
grow towards the cytosol
rich in motor proteins and are anchored to the integrins allowing filaments to move forward
at trailing edge of cell integrins internalized and recycles and stress filaments are disassembles
cell cycle checkpoints
checkpoints control transitions between phases to avoid unnecessary energy waste
called cyckin dependent kinases (CDKs)
CDKs bind to respective cyclins to become active which then phosphorylate other proteins to trigger the next stage of the cycle
CDK + associated cyclin
G1 to S
- CDK 4 + cyclin D
commits to replication (G1)
- CDK 2 + cyclin E
initiates replication (S)
- CDK 2 + cyclin A
promotes mitosis (G2)
- CDK 1 + cyclin B
cell cycle phase order
G1 phase
G0 phase
G1/S checkpoint
S phase
S/G2 checkpoint
G2 phase
G2/M checkpoint
M phase
mitotic spindle checkpoint
G1 phase
gap 1
cell active and growing
not committed to undergoing division
G0 phase
gap 0
technically not apart of cycle
when cell is resting (nerve/muscles cells)
G1/S checkpoint
cell proteins check for DNA damage
start point, commits cell to progression through cycle
activates signals allowing cell to divide
S phase
cell replicates entire genome
centrosome duplicated
S/G2 checkpoint
DNA integrity checked
G2 phase
last chance for cell to grow before division
cytoplasm and cellular contents (endomembrane system) increased in preparation for division
G2/M checkpoint
large scale rearrangement to structure of cell
increase in cell volume causes progression through checkpoint
M phase (mitosis)
division occurs
mitotic spindle checkpoint
ensure all chromosomes are properly separated preventing chromosome imbalance
ensures cytokinesis only occurs successful completion of mitois
p53 protein
tumour suppressor proteins that ensures cells with damaged DNA dont divide
initiate apoptosis in cells with damaged DNA
if dysfunctional cells can evade apoptosis and replicate uncontrollable
phases of mitosis
- interphase (not a phase)
- prophase
- prometaphase
- metaphase
- anaphase
- telophase
- cytokinesis (not a phase)
interphase
when DNA replication occurs (G1, S and G2 phase)
prophase
chromosomes condense and pack into chromatids
sister chromatids connected at centromere
gene transcription stops
endomembrane system dissolves into tiny vesicles, mitochondria remain but are randomly distributed in cell
nuclear envelope dissolves, choromsomes released into cytosol
mitotic spindle forms around each centrosome
centrioles form (MTOCs)
prometaphase
kinetochore forms (complex that binds to chromatids, each sister chromatid has two, one on each side)
kinetochores use ATP to polymerize and depolymerize microtubule spindle fibers allowing chromosomes to move to the center of the cell
metaphase
all chromosomes arrive at spindle equator
chromosomes successfully attach to kinetochore microtubule which now pulling equally in both directions
mitotic spindle checkpoint occurs ensure proper alignment at equator
anaphase
proteins binding sister chromatids cleaved, kinetochore microtubules shorten
in last anaphase chromosomes reach maximum condensation level
additional microtubule organize around spindle equator in preparation for cytokinesis
telophase
reorganization of cell
nuclear membrane reform around chromosomes
interphase cytoskeleton and endomembrane system reform
cytokinesis
animal cell, contractile ring forms where spindle equator was
ring tightens cell divides in half, moment when plasma membrane snaps (hydrophobic nature cause rupture to reseal spontaneously)
after occurs two new cells enter interphase
reform junctions and interior of normal cell
