Actin based cell movements Flashcards
(29 cards)
Describe actin filaments
Linear polymers of a single globular protein - G-actin
F-actin (filamentous actin) microfilaments are helical polymers
A left handed helix with a rotation of of 166 degrees per subunit
There are 13 monomers per helix repeat of 37 nm
at 7nm in diameter, they are flexible ropes
Talk about the G-actin monomer
Monomeric G-actin binds one ATP
G-actin hydrolyses ATP
Subdomain 2-4 surface binds to 1-3 surface, resulting in filament polarity
the monomer has different protein surfaces exposed
Describe the dynamics of actin filaments
The plus end has the highest binding affinity to ATP-binding G-actin
After polymerisation into microfilaments, actin monomers hydrolyse their bound ATP which destabilises the filament
ADP-binding subunit dissociation from minus end
Treadmilling - Actin elongates at plus end and shrinks at minus end
the overall filament will move in the direction of the plus end
What are the 5 functions of actin binding proteins
Actin binding proteins determine the rate of filament assembly and stability
Nucleation
Capping
Severing - cut the filament to smaller parts, exposing ADP bound actin leading to depolymerisation
Sequestering - cross link to make larger structures
Bundling
What is the function of the microfilament network***
Strong in tension, weak in compression (more useful in pulling than pushing)
1 - Linear pathways for organelle movement in plants and fungi
2 - in animals, they form contractile systems, together with motor proteins (in plants cell these molecules are not needed)
3 - When cross-linked, they can have a variety of structural roles, and can push the growing margins of the animal cell forward
What are myosin motor proteins
Myosins are a family of motor protein
Two catalytic ATPase head walk along actin filament
the motor heads can convert chemical energy released by the hydrolysis of ATP into mechanical movement
The motor domain is connected a neck domain to a tail domain which interacts with cargo or dimerises
the myosin cross bridge cycle
The myosin head undergoes dramatic changes in conformation depending on its binding:
Pi release causes strong filament binding and conformational change in neck region
Power stroke
the distance moved by each myosin head is 6nm
heads are detached most of the time
Organelle and vesicle transport in plants
How does cytoplasmic streaming help with diffusion?
In plants, small organelles and vesicles are continually moved around the cytoplasm - the drag caused by moving organelles cause the cytoplasm to cycle around the cell leading to cytoplasmic streaming
movement is powered on actin filaments, using myosin motor proteins
it is used to overcome diffusion barriers in extremely large vacuolate plant cells
In animals, how do actin and myosin II form contractile arrays
clue: myoblast, myotube, part of muscle fibre
Muscles fibres are giant animal multinucleate syncytial cells, about 50 micrometers in diameter, formed by the fusion of mono nucleated precursors, myoblasts .
within each fibre are many myofibrils , forming the contractile apparatus.
myoblast –> myotube –> part of muscle fibre
Describe sarcomere structure
Each myofibril is a highly organised linear array of sarcomeres, the contractile units
Actin-containing thin filaments project with opposite polarities from the two Z discs. the actin plus ends are embedded in the Z discs
interdigitating between the thin filaments are thick filaments made of the motor protein myosin
Describe myosin thick filaments
actin is the thin filament
muscle myosin spontaneously assembles into bipolar thick filaments , with a bare central zone
Elsewhere, myosin heads project sideways from the thick filaments, with three fold symmetry and a period of 43nm, the heads projecting in nine radial positions
The sliding filament model of contractions (relies on sliding not polymerisation)
Shortening of the sarcomere is brought about by two sliding of two filament sets, without change in filament lengths
This shortening is caused by myosin molecules binding to the actin filaments and pulling them towards the centre of the sarcomere - actin is in tension –>
Filaments are of defined lengths
Name the giant proteins that act as molecular rulers
Titin and nebulin control the lengths of thick and thin filaments
Titin’s elastic ends also holds the myosin thick filaments in the centre of the sarcomere
Myosin cross-bridge binding
How does the spacing of myosin heads maintain tension ?
The spacing of the myosin heads is out of register with the spacing of the myosin binding sites on the actin thin filament.
Repeat distance of 129nm of myosin heads
each head 6nm per cycle
Repeat distance of myosin binding sites are 37nm
Furthermore,
the symmetry of the myosin heads and their surrounding actin sites differ.
The myosin heads stick out in NINE radial positions
Each thick filament it is surrounded by SIX actin filaments
Therefore,
Myosin heads cannot all bind actin at the same time, ensuring some heads are attached at ALL times to maintain tension
How are actin-myosin interactions coordinated?
The thin filament contains accessory proteins troponin and tropomyosin
Two calcium ions bind to troponin C
The troponin complex changes shape and induces tropomyosin to roll away from the myosin binding site on the actin microfilament
Myosin can now bind to the actin filament and cause contraction
Contraction will continue as long as calcium ions are present
How does calcium release co-ordinate contraction
Calcium is released from stores in the sarcoplasmic reticulum
Conc of Ca in SR is about 10,000 times greater than the resting concentration in the muscle cytoplasm.
The SR membrane is in electrical contact with the plasma membrane
Arrival of action potential opens calcium channels in the SR increasing ca2+ concentration about 10 fold.
How does cytokinesis work in animals and fungi?
Cell separation after mitosis is brought about by a contractile ring of actin and myosin II
Form a ring of mini sarcomere like structures .
Actin filaments pushed together by myosin into ring structure – > pinch the cell.
How does microfilament organisation work in non muscle cells?
Cytoplasmic bundles of F actin and myosin II form stress fibres - keep the shape of the cell structure
Stress fibres are anchored at one end at the PM (focal adhesion point) and at the other either in a peri-nuclear cage or PM
Focal adhesion points anchor cells to other cells or extracellular matrix .
stress fibres are contractile actin arrays
What is the structure of a stress fibre?
Stress fibres contain both bipolar actin filaments and myosin II mini-filaments and are contractile
Mini-sarcomeres
Actin polymerisation can drive rapid movement in cell migration (not myosin movement!) . Given an example
Keratocytes from fish scales move very fast
How does microfilament organisation in crawling cells work
Lamellipodium
Filopodium
Stress fibres
Lamellipodium - large, flattened cell extension at the leading edge, quasi-2D branched actin meshwork – pulls the cell forward
Filopodium - small, dynamics cell projections, actin bundle, senses environmental signals
Stress fibres - contractile bundles, push the trailing edge
Describe in more detail the branched actin filament array at the leading edge e.g. in fish scale keratocyte cell
The lamellipodia of a fish scale keratocyte cell contains a dense array of short actin filaments
The short filaments in these dense networks provide enough rigidity to push on the membrane at the leading edge
These show characteristic 70 degree angle intersections.
What are ARPs - actin related proteins
ARP 2/3 complex can bind to pre-existing actin filaments at a 70 degree angle
Nucleate formation of new filaments creating a cross-linked meshwork threading lamellipodia of motile cells.
How are ARPs similar to G-actin?
ARPs cannot participate in F-actin filament but can bind actin minus ends.
In vivo the ARP 2/3 complex nucleates actin filament, capping the minus-end
analogous to gamma tubulin nucleating MT
