Cytoskeleton

Question 1

What is cytoskeleton? What are its functions?

Answer 1

= internal network of protein filaments that span over cytoplasm
- Functions:
- structure and support (especially useful for animal cells without cell wall)
- dynamic movements e.g. contraction, crawling
- changes in shape e.g. during embryonic development
- builds transport system between organelles
- segregation of chromosomes during cell division

Question 2

What builds up the cytoskeleton?

Answer 2

Three types of protein filaments:
1) Intermediate filaments - specific family of fibrous proteins
2) Microtubules (myosin filaments) - globular tubulin subunits
3) Actin filaments - globular actin subunits

Question 3

What characterizes Intermediate filaments?

Answer 3

• tensive strength, most durable, toughest
• function: enable cells to survive mechanical pressure (e.g.from being stretched)
• named “intermediate” because in muscle cells (where they’ve been discovered) their diameter falls between actin and myosin filaments

Question 4

Where do we find intermediate filaments?

Answer 4

• In most animal cells -> network surrounding nucleus and going into periphery
  
  • anchored to the plasma membrane at cell-cell junctions = desmosomes
• Also in nucleus -> form a tough meshwork = nuclear lamina, which underlies and supports nuclear envelope

Question 5

What makes intermediate filaments so strong?

Answer 5

• it is a rope of multiple fibrous strands (filaments) twisted together
  
  • extended alpha-helix forms a dimer with another aloha-helix => coiled-coil configuration
    -> two dimers running in opposite directions get twisted = tetramers
    -> tetramers associated with each other side-by-side = filaments

Question 6

What differentiates intermediate filaments from the actin ones?

Answer 6

• The dimers are in opposite directions -> the ends will be the same
  -> once filaments assembled their ends are also the same
  -> they rely on non-covalent bonding along the length of the protein (-> provides the strength)
• BUT actin filaments microtubules require polarity for their function (i.e. different ends)

Question 7

Are all intermediate filament proteins the same?

Answer 7

Yes and no
- the central domain of each rod is the same (amino acid sequance, size, diameter)
- BUT the terminal domains differ from one type to another

Question 8

What kind of cells have abundance of intermediate filaments? How does it help them?

Answer 8

E.g. along axons of nerve cells, muscle and epithelial cells
- they distribute the mechanical forces applied on these cells to keep them from tearing

Question 9

What classes of intermediate filaments do we have? And what cells do they belong to?

Answer 9

1) Keratin filaments in epithelial cells
2) Vimentin and vimentin-related filaments in connective-tissue cells, muscle and glial cells
3) Neurofilaments in nerve cells
4) Nuclear lamina in nuclear envelope

Question 10

What is the most diverse class of intermediate filaments? What characterizes them?

Answer 10

Keratin filaments - present in epithelial cellsof the tongue, skin, gut, cornea, hair, feathers -> need tohave types adjusted to the special needs of these organs

• typically a mixture of different keratin proteins
• spans the cell from one side to the other
• some else connect adjustent epithelial cells via their desmosomes -> creates epithelial sheets that can further distribute mechanical pressure when the skin gets stretched

Question 11

How is epidermolysis bullosa simplex connected to intermediate filaments?

Answer 11

= mutation in keratin genes that interfere with the proper formation of keratin filaments
-> skin is highly vulnerable to mechanical injury (even just small pressure can result in rupturing of the cells -> causing blisters)

Question 12

What further stabilizes the intermediate filaments?

Answer 12

• Accessory proteins e.g. Plectin
  
  • cross-links the filaments into bundles, links themto microtubules and actin filaments

Question 13

How do intermediate filaments of the nucleus differ from those of cytosol?
What happens during cell division?

Answer 13

• Instead of rope-like structure they form two-dimensional meshwork
• constructed from a specific class of proteins = lamins
• during cell division lamins disassembles and reforms -> controlled by phosphorylation and dephosphorylation
  - Phosphorylated by protein kinases -> conformation change weakens binding between lamin tetramers -> falling apart
  - Dephosphorylated by protein phosphatases helps with building it back up

Question 14

Which disease is connected to nuclear lamin?

Answer 14

Progeria = rare disorder causing children to age prematurely e.g. wrinkled skin, hair or teeth loss, cardiovascular diseases

• Not yet know how loss of nuclear lamin could cause this - but likely due to impaired cell division, excess of cell death

Question 15

What characterizes microtubules?

Answer 15

• Long, hollow tubes
• Can desassemble in one location and assemble in another
• Grow from centrosomes to periphery
• Function:
  
  • provide tracks for transport of vesicles, organelles, and other molecules

Question 16

What forms of microtubules can you think of?

Answer 16

• Mitotic spindles
  = assembly of microtubules during mitosis which helps to segragate chromosomes equally into the daughter cells
• Cilia and flagella
  = structures that extend over the surface of the cell and enable e.g. movement

Question 17

How are microtubules build?

Answer 17

• from subunit molecules = tubulins
  
  • alpha-tubulin monomer binds noncovalently to beta-tubulin monomer -> forming tubulin dimer
    -> stack together into a hollow cylindrical shape
    -> the “cylinders” = protofilaments are stacked parallel to one another
    - each has structural polarity (alpha-tubulin always on one side while beta-tubulin always on the other) -> microtubule has beta-tubulin end = plus end and alpha-tubulin end = minus end

Question 18

What are the centers organizing microtubules in animal cells?

Answer 18

Centrosomes
- found close to the cell’s nucleus
- consists of pair of centrioles perpendicular to one another + surrounded by matrix of proteins
- the matrix consists of many ring structures formed by gamma-tubulin -> each ring complex forms a starting point of microtubule = nucleation
- minus end starts at centrosome -> micrutubule grows towards the plus end

NOTE: gamma-tubulin provide ring sites to which alpha or beta can bind - as it would be diffucult to start the initial ring themselves

Question 19

What do we mean by “dynamic instability”?

Answer 19

= the concept of polymerazation and dypolymerazation of microtubules depending on the cel’s needs
- centrosome is continuously creating knew microtubule bridges
-> if plus end reaches its destination, the microtubule is prevented from deassembling
-> if it doesn’t = destroyed

Question 20

How is the dynamic instability driven?

Answer 20

Hydrolysis of GTP
- Each free tubulin dimer contains one GTP molecule bound to beta-tubulin -> binds to the growing microtubule -> GTP hydrolyzes into GDP
- BUT the hydrolysis is slower than the adding of dimers -> the growing part of microtubules will be composed of GTP-tubulin dimers (“GTP cap”) which bind next/or previous dimers more strongly -> growing continues

• If GTP is hydrolysed earlier (could be just by randomness) the part will become GDP-tubulin
  - less association with other dimers -> diassemling is prefered (-> might even eventually disappear)

Question 21

How may microtubules dynamics be used in cancer treatment?

Answer 21

Since in the caseof cancer there tend to be an abnormally frequent division of cells (which uses mitotic spindles to divide its chromosomes) - drugs acting on microtubules could preferentially kill of these cells
- E.g. Colchicine binds to free tubulin in cytosol and prevents it from linking to the growing microtubules -> disassembly will be favoured
- E.g. Taxol binds to microtubules and prevents them from loosing subunits (can grow but cannot shrink)

Question 22

True or False: “Dynamic instability may happen at different rates”

Answer 22

True
- During mitosis the rate of DI is much higher -> enables rapid disassembly of microtubules and formation of mitotic spindles

• After cell differentiates -> cytosolic microtubules are slower at this
  
  • already stabilized by proteins binding to either side of the microtubules -> maintaning organization of specialized cells

NOTE: spcialized cells tend to be polarized
- e.g. plus end being at the terminal buttons of an axon -> helps with trafficing of vesicles (way faster than just free diffusion)

Question 23

What other molecules help microtubules out?

Answer 23

Microtubule-associated proteins = variety of accessory proteins that may
- stabilize against disassembly
- link other microtubules and filaments
- transport as motor proteins

Question 24

Why does it seem like some organelles and vesicles seem to be moving in “small, jerky” steps? What is it called?

Answer 24

• This is called saltatory movement - and is more sustainable and directional than random thermal motion
• Driven by motor proteins that function on the basis of repeated cycles of ATP hydrolysis

Question 25

There are different motor proteins - that bind todifferent filaments, go in different directions and carry different cargo -> name 2 known families.

Answer 25

- Kinesin - move towards the plus end of microtubules (outwards from the cell body e.g. terminal buttons) - Dyneins - move towards the minus end (towards the cell body) - Both are dimers with 2 globular ATP-binding heads and a single tail - heads attach it to the microtubules, bind ATP and lead hydrolysis (binding -> hydrolysis -> release -> binding) - tail binds a specific cargo

Question 26

What is meant by "motor proteins help position organnelles in space"?

Answer 26

E.g. as cells grow kinesin attaches to the outside of ER and stretches outward (from the nucleus) X dyneins stretch Golgi apparatus inward towards the nucleus NOTE: if cell is treated by colchicine causing microtubules to disassemble -> ER and Golgi apparatus collapses

Question 27

What are cilia and flagella?

Answer 27

Cilia are hairlike structures that propel single cells through fluid - could be used for locomotion or collections of food - e.g. epithelial cells lining the human respiratory tract -> sweap mucus containing trapped dust particles towards the throat -> swallowing and eventually eliminating from the body Flagella - longer, meant to move the cell itself rather than something on its surface

Question 28

What generates the movement of cilia and flagella ?

Answer 28

Produced by bending of the core as the microtubules slide against each other - also use ciiary dynein attached to the tail of one microtubule -> while its two heads interact with the adjacent microtubule to generate a sliding force between the two microtubule - due to these links it becomes bending (rather than actual sliding)