Lecture 8- Microfilaments

Question 1

Cytoskeleton is composed of

Answer 1

Microfilaments
Intermediate filaments
Microtubules

Question 2

Actin structure and assembly

Answer 2

Microfilaments
ATP and Mg are cofactors
ATP hydrolyzed to ADP after subunits are assembled into the filament

Question 3

Filamentous actin (structure and assembly)

Answer 3

Self assembly of actin monomers into two stranded 5-7 nm diameter helical filaments of variable length.
Uniform orientation of monomers gives a structural polarity of filaments

Question 4

Treadmilling of Actin

Answer 4

Plus end is fast growing
Minus end appears to shrink
Hydrolysis of ATP to ADP on actin subunits of a filament results in different assembly equilibria at plus and minus ends

Question 5

Actin binding proteins

Answer 5

Two major classes: Thymosins and profilins
Can generate force (myosin), regulate motility (troponin), crosslink (filamin), bundle (alpha actin and fimbrin), cap filaments (alpha/beta capping protein), anchor filaments to other structures such as the plasma membrane (talin)

Question 6

Cell cortex

Answer 6

Combination of the plasma membrane and an underlying mesh of crosslinked actin filaments

Question 7

Actin associated membrane structures description

Answer 7

May be involved in contraction, extension of the plasma membrane to form filopodia and lamellipodia, and stabilization of a particular membrane shape

Question 8

Actin associated membrane structures list

Answer 8

• Microvilli–ex: absorptive cells of the intestines
• Stress fibers–antiparallel, contractile bundles of actin filaments that terminate on the plasma membrane at focal contacts
• Lamellipodia–extensions of the cell membrane in a migrating cell
• Contractile ring of a dividing cell

Question 9

Lamellipodia formation

Answer 9

ARP (actin related protein) complex mediates assembly of a branched array of actin filaments at the leading edge of an extending cell membrane

Ex: fibroblasts migrating through the extracellular matrix in connective tissue

Question 10

Phalloidin & Cytochalasins

Answer 10

• Anti actin drug
• Stabilizes actin filaments by binding along the sides of the filament
• Disrupts cell motility and cytokinesis

Question 11

Myosin II structures

Answer 11

Assembles into bi-polar filaments with heads that face two directions, interact with two overlapping sets of actin filaments polarized in opposite directions

Question 12

Myosin I structures

Answer 12

Do not form filaments

Walk along actin filaments toward their plus end

Question 13

Cell motility by coordination of myosin and actin assembly/disassembly

Answer 13

• Actin assembly extends the lamellipodium
• Attachment to the substrate and contraction of microfilament bundles pulls the center of the cell forward
• Detachment of the back end of the cell and further contraction brings the back end of the cell forward

Question 14

Regulation of Actin-Myosin contraction in non-muscle cells

Answer 14

• Phosphorylation of myosin-associated proteins called myosin light chains
• Ca concentration changes cause phosphorylation of light chains by activating myosin light chain kinase
• Called myosin based or thick filament based regulation

Question 15

Skeletal muscle structure

Answer 15

• Long multi-nucleate cells containing many long myofibrils
• Divided into many short repeating units called sarcomeres (z line to z line)
• Plus ends of actin filaments attach to z disks, minus ends extend away but do not overlap in center
• Bipolar myosin filaments held in the middle of each sarcomere
• Myofibrils bound to each other by desmin intermediate filaments
• Entire array anchored to plasma membrane by many proteins one of which is dystrophin

Question 16

Mechanism of muscle contraction

Answer 16

1. Rigor– myosin head is tightly bound to the actin filament and nucleotide free
2. Release– ATP binding to the myosin head lowers the affinity of myosin for actin resulting in release
3. Cocked–ATP hydrolysis causes a 5nm translocation of the head to cock it in preparation for the power stroke. This conformation has weak affinity for the actin filament.
4. Force generating–Dissociation of the inorganic phosphate increases the affinity of the myosin head for the actin filament and activates the power stroke.
5. Attached–Dissociation of ADP is stimulated by translocation of the myosin head back to its original conformation.

Question 17

Troponin-tropomyosin complex

Answer 17

• Involved in regulation of skeletal muscle contraction
• Called actin or thin filament based regulation
• In the absence of calcium tropomyosin will block the myosin binding site
• Binding of calcium to troponin C causes troponin I to release its hold on actin thus allowing tropomyosin to move away from the myosin binding site and allow myosin to bind to actin.