7.3 Microfilaments II

Question 1

Protein that is going to organize actin filaments has to have how many domains? Why does this make sense?

Answer 1

2 actin-binding domains.

This makes sense b/c if a protein is going to organize actin filaments it has to have more than one binding site so that it can hold onto more than one filament

Question 2

How would short actin binding proteins be used by the cell?

Answer 2

To create a very densely packed actin filament bundle.

Ex: fimbrim and villin

Question 3

What are the different types of actin bundling proteins that regulate organization of actin filaments?

Answer 3

Fimbrin + villing (they are short monomers -tight bundles)

Alpha actinin (dimer lose bundles in smooth and cardiac muscle)

filamin (y shaped, forms cross link between existing actin filaments creating meshwork)

Spectrin + Myosin I (attach filament to cell membrane, spectrin in red blood cells and myosin does the same but to microvilli)

Question 4

Which structure contains alpha actinin?

Answer 4

Cardiac muscle cells
Smooth muscle
Muscle fibers and focal adhesions

Question 5

What does alpha actinin do?

Answer 5

Forms loose actin bundles
It is also part of actin filament arrays in all types of muscle cells which allows myosin to interact with actin filaments.

Question 6

What are microvilli?

Answer 6

Tiny hair-like protrusions found on the top of cells in the intestine, kidney, and other places where fluid and dissolved nutrients are absorbed. Their role is to greatly increase the cell surface area for absorption.

Question 7

What protein supports microvilli?

Answer 7

By small tightly packed bulges of cross-linked actin filament. Villin and fimbirn

Question 8

How does filamin work? What happens if filamin is mutated?

Answer 8

filamin forms actin meshwork by crosslinking existing filaments. it is s a Y-shaped cross-linking protein and it cross-links actin filaments into 3-dimensional meshwork.

Cells that have a filamin mutation have trouble keeping their plasma membranes under control and their membrane bubles away from the cell surface.

Question 9

What is One of the places actin meshworks is important (aka filamin)?

Answer 9

The layer of actin filaments found just under the plasma membrane that gives the cell membrane its strength.

Question 10

Which proteins attach the actin cytoskeleton to cell membranes?

Answer 10

Spectrin - blood

Myosin I - microvilli

Question 11

What is the function of spectrin? Where is it found?

Answer 11

Found in red blood cells ghost

Spectrin links the membrane of blood cells to an actin meshwork. (links the actin filament system to proteins that are embedded in the cell membrane and this forms the structural shell of a red blood cell that the membrane is then stretched over.)

Question 12

What is the role of myosin I?

Answer 12

Myosin has the same role as spectrin but instead of attaching the membrane to red blood cells it does it to microvilli.

It is a motor protein that links membranes to actin in microvilli
Myosin are modified to bind membranes with their tail domains and then attach the membranes to the actin filament system with their motor-head domains.

Question 13

Summary of Actin Accessory Proteins

Answer 13

Nucleation: Profilin promotes, Thymosin inhibits by sequestering, Arp2/3 creates 3D meshwork

Disassembly: Gelsolin (Ca++ activated) severs, cofilin destabilizes

Stabilization: CapZ stabilizes

Bundling: Alpha-actinin (loose), Fimbrin, Villin (tight)

Meshworks: Filamin

Membrane attachment: Spectrin (blood cells), Mysoin I (microvilli)

Question 14

What are lamellar protrusions?

Answer 14

Flat sheets of cell membrane and thin layers of cytoplasm that protrude at the edge of a cell

The formation of lamellar protrusions are coordinated by several types of actin associated proteins at the same time (Capz, Arp2/3, and cofilin)

Question 15

What happens if lamellar protrusions make an attachment to the cell substrate?

Answer 15

They will become a new cell extension and the cell can flow into them

Question 16

What happens if lamellar protrusions don’t attach to anything?

Answer 16

If they don’t attach to anything, they will often migrate backward across the top of the cell and then remerge with the cell body.

Question 17

Where are lamellar protrusions normally found and why?

Answer 17

Front of a moving cell b/c cells can move into them

Question 18

What is going on at the front of a lamellum?

Answer 18

1) uncapping (capz)
2) Assembly (Arp 2/3)
3) Disassembly (Cofilin)

Inside Arp2/3 nucleation creates a 3dimensional meshwork of actin.

Many of the actin filaments ends are capped with capZ.
When the capping protein is deregulated, new actin assembly occurs in that area and it pushes the cell membrane forward.

A key issue is that the growth of actin filaments could quickly deplete the concentration of G-actin in that area and assembly would stop but to deal with this Cofilin enters into lamellar to promote disassembly of older filaments. This then provides new subunits that can be used for additional rounds of assembly.

Question 19

In lamellar protrusions, the growth of actin filaments could quickly deplete the concentration of g-actin in that area and then assembly would stop. How is this issue solved?

Answer 19

Cofilin enters into lamellar protrusions just behind the wave of assembly and it promotes the disassembly of older filaments. This provides new subunits that can be used for additional round of assembly

Question 20

What are platelets?

Answer 20

Found in the bloodstream they are membrane bound structures that are smaller than red blood cells and are not involved in carrying oxygen. Instead they play a role in clot formation.

Question 21

What happens when platelets are inactive?

Answer 21

they are small lentil-shaped structures that move easily in the bloodstream

Question 22

What happens when platelets are active?

Answer 22

when they are activated by a clotting signal, they form a sort of spider-shaped structure that can easily plug holes in blood vessels Gelsolin plays a role in activating.

Question 23

What are the sequence of events that cause the active and inactive state of platelets?

Answer 23

1) in an inactive platelet, there are a small number of long, capped actin filaments.
2) When the platelets are activated, there is an influx of calcium that activates gelsolin. Gelsolin then cuts up the existing actin filaments creating many new nucleation sites.
3) the rising concentration of calcium in platelets is transient and when the concentration of calcium drops again, the gelsolin is inactivated and extensive actin polymerization occurs

3) the actin filaments that grow are then organized by the activities of actin-bundling proteins like fimbrin, α-actining, and filamin and that gives platelets a spider-shape
These events occur sequentially!

Question 24

What global regulator decide whether a cell is going to be highly motile or highly stationary or something in between?

Answer 24

Ras family members Rac, Rho and CDC42 are cell signaling proteins that coordinate actin assemblies.

Question 25

What happens when there’s a single mutation in the gene for Ras?

Answer 25

Causes a cell to become cancerous

Question 26

Who are the Ras family members?

Answer 26

Rho- form strong attachment
Rac -move rapidla/lamellar protrustions
CDC42 - philopodia

Question 27

What does activated Rho do?

Answer 27

causes the cells to create strong cables of actin filaments, called stress fibers, as well as many vinculin-containing structures all around the cell.

Highly focused to their attachment site

Question 28

When will a cell active Rho?

Answer 28

Rho is activated in cells that want to contract or to attach strongly to the surface they are on

Question 29

What does activated Rac do?

Answer 29

Rac causes the formation of lamellar protrusions

When is Rac activated?

When cells want to move rapidly

Question 30

What is the role of CDC42 ?

Answer 30

causes the formation of thin, finger-like protrusions of the cell which are called philopodia

Question 31

How do philopodia differ from microvilli?

Answer 31

Philopodia are different from microvilli because they are unstable and they can form surface attachments which contain vinculin. However, many of the same actin bundling proteins are involved in their formation. Some cells use philopodia to explore their environment before deciding where to move.

Question 32

Summary of Actin Regulation:

Answer 32

Complex behavior requires coordination of assembly, disassembly and organization: Lamellar protrusions: CapZ, Arp2/3, Cofilin Platelet activation: Gelsolin, Fimbrin, Filamin

Actin supported behaviors are regulated by cell signaling processes Rac, Rho, cdc42 are master controllers of actin filament organization.

Question 33

What is a keratocyte cell?

Answer 33

a single, large lamellar protrusion which drags the cell body around behind it. the actin filaments are found at the very edge of the cell where they are pushing the membrane forward while cofilin is found in a zone that is just behind the area of assembly but it is excluded from that front zone.

these cells are a favorite model for people interested in studying actin
assembly

Question 34

How can regulation of actin filament assembly be studied?

Answer 34

Keratocyte cells

Platelete activation

Question 35

When is rac activated?

Answer 35

When cell wants to move rapidly. Rememeber rac creates lamellar protrusions, which are in the direction that the cell moves