Transcript Cytoskeleton

Cytoskeleton
A cytoplasmic system of fibers -> critical to cell
motility (movement)
Macrophage cytoskeleton
Cytoskeleton of a lung cell in mitosis
1
The cytosol:
20-30 w% of cytosol are proteins -> ¼ - ½ of total
protein is in cytosol
Protein conc. 200-400 mg/ml -> complexes of protein
It is believed that cytosol is highly organized
-> Most soluble proteins are
- bound to filaments
- localized in specific regions
2
Cytoskeleton of an epithelial cell and a migrating cell
3
Eukaryotes
Bacteria
Actin filaments (AF)
Intermediate filaments (IF)
microtubule (MT)
4
Cytoskeleton is made out of 3 different types of filaments
5
Filaments differ in Size, Shape and Flexibility
6
Filament network in the cell
Microtubules network
Starting from the MT center near nucleus
Filament network (fluorescence)
From the nucleus to the plasma membrane
7
Cytoskeleton supporting the plasma membrane in human red blood cells
8
Cell Signaling Regulates Cytoskeleton Function
9
10
1. Microfilaments and Actin Structures
Actin cytoskelet in a moving cell
11
1. Microfilaments and Actin Structures
Actin monomers assemble into long helical polymers with polarity
12
Actin Filament Assembly
13
Actin Filaments grow faster at (+) end than at (-) end
The rate of addition of ATP-G-actin is much faster at the (+) end than at the (-) end
(rate of dissociation is similar) -> lower critical concentration (Cc) at (+) end
in steady state -> filament grows preferentially at the (+) end
If actin conc. is between Cc- and Cc+ (steady
state) -> actin subunits flow through
filaments by attaching to (+) end and
dissociating from (-) end
Treadmilling phenomen -> involved in
movement of cells
14
Capping Proteins Block Assembly and Disassembly at Actin Filament Ends
The presence of these 2 proteins at opposite ends prevent actin from dissociating during muscle contraction
15
Actin Filament Branching
Nucleation of
branching
mediated by
Arp2/3
16
Movement of invaders inside the cell
Most infections are spread by lysed cells.
Some Bacteria (Listeria monocytogenes) or viruses (vaccinia – related to smallbox virus) escape from
cell on the end of a polymerizing actin filament.
These organisms or viruses move through the cytosol at rates of 11μm/min.
Actin generates the force necessary for movement
17
Proteins that organize Microfilaments into networks
Forms bundles
Filamin forms networks
18
Filaments attached to Membranes
Microvilli on an epithelial cell showing polarity of actin filaments
19
Myosins - Cellular Motor Proteins
Tail:
-> Locatized to cellular membranes
-> vesicle attached (cargo)
Form thick filaments in muscles
S1 motor domain
Head -> Motor domain (S1) -> ATP depentend
20
Myosin heads walk along actin filaments -> towards (+) end
Sliding-filament assay:
Myosin tail absorbed onto glass
surface -> a solution of actin
filaments allowed to flow
through
In presence of ATP myosin
heads walk towards (+) end of
actin filaments -> sliding of
filaments
-> Movement of labeled
actin filaments
21
Myosins – Motor proteins responsible for cell movement
These are the most important 3 myosins (out of ca. 40 we have in humans)
Loss of more specialized ones -> causes deafness/blindness
22
Myosin motion along actin
Length of the neck domain ->
determines rate of movement
Step size
-> Moves in 72 nm steps
23
Actin fibers in the muscle
24
Skeletal muscle contraction is regulated by Calcium and actin binding proteins
Tropomyosin (TM)
Troponin (TN)
25
Cell Locomotion
Coordination of motions generated by different parts of the cell
Movement of fish epidermal cell
Cell locomotion mechanism:
Includes actin polymerization and branchinggenerated movement at the edge, assembly of
adhesion structures, and contractions
mediated by myosin II
26
Myosin V Carries Many Cargoes
Myosin V:
-> carries secretory vesicles, organelles,...
-> Used to prepare nucleus for mitosis
-> used to segregate organelles
27
2. Intermediate Filaments
IF differ in stability, size and structure from other
cytoskeleton fibers:
- IF are extremely stable (hair, nails, wool)
-10 nm diameter
- α-helical rods -> assemble into ropelike filaments
- assemble from different IF proteins
- assembly through several intermediate structures
Keratin and lamin IF
Intermediate structures in the assembly of IF
28
29
Cross-links between Microtubules and
Intermediate Filaments in Fibrioblast cells
Microtubules (red), Intermediate Filaments (blue),
connection between fibers (green)
30
3. Microtubules
Kinesin-powered movement of a
vesicle along a microtubule
Microtubules are involved in cell
movement:
- Beating of cilia and flagella
- transport of vesicles in the
cytoplasm
Microtubules organizing center (MTOC)
31
Microtubules organization
2 type of MT in cells:
- Stable and long-lived (found in non-replication cells) -> in cilia, flagella, neurons
- unstable and short-lived (found in mitosis) -> spindle-shaped apparatus that partitions chromosomes
equally to daughter cells
32
Microtubules Arrangement
Flagella
33
Microtubules assemble from Microtubule Organizing Centers (MTOCs)
MTOCs in non-mitotic cells -> centrosomes
34
Microtubules Assembly preferably at (+) end
Nucleation of microtubule assembly
-> Treadmilling
35
Microtubules Assembly/Disassembly
Colchicine and Taxol:
Drugs that interfere with Microtubules
Assembly/Disassambly
Colchicine: 2500 years ago Egyptians treated heart
problems
Nowadys: treatment of gout, skin and joint diseases
Taxol: (stabilizes Microtubules)
Anticancer agents -> treatment of ovarian cancer
36
Microtubules Dynamic Instability
Presence of GTP-β-tubulin cap
determines stability
37
38
Kinesin and Dynein – two Families of Motor Proteins
Responsible for Transport along Microtubules
Microtubules based vesicle transport
39
40
Kinesin-catalysed Vesicle Transport
Carries cargo
41
Kinesin-1 uses ATP to walk down a microtubule to the (+) end
42
Dynein-catalysed Vesicle Transport
Moves towards the (-) end of microtubules
43
Microtubule motors in a Cell
Kinesins -> transport to cell periphery (+)
Dyneins -> transport to cell center (-)
44
Cooperation of Myosin and Kinesin at the cell cortex
Secretory vesicles are handed over
from Kinesin to Myosin -> last part of
secretory pathway
45
Rotory Motors – Cilia and Flagellar
Bacterial Flagella (E. coli, Salmonella)
Rotation of Flagellar -> Motion
Sperm
46
The Flagellar Motor
47
Bacterial Flagellum is made out of subunits
48
Motion of E. coli
The points show the locations of
the bacterium at 80 ms intervals.
Changing of direction: Tumbling
is caused by an abrupt reversal
of the fagellar motor
A second reversal restores
smooth swimming -> almost
always in a different direction
49
Reversal of the direction of Flagellar Rotation is obtained by
Proton Transport through Motor
Proton flow drives flagella rotation !!!
50
Chemotaxis Signaling Pathway
Receptor in plasma membrane
initiate signal pathway ->
Phosphorylation of CheY protein
-> P-CheY binds to flagellar
motor -> clockwise rotation
favored
Attractant binds to receptor ->
pathway blocked -> smooth
swimming
Repellant binds to receptor ->
pathway stimulated -> more
frequent clockwise rotation ->
tumbling
Direction of bacterial movement depends on chemical substances:
-> Bacteria swim towards high concentrations of glucose - chemoattractans
-> Bacteria swim away from harmful substances, such as phenol - chemorepellants
51