Cytoskeleton and Cell Junctions

Question 1

cytoskeleton functions

Answer 1

=large dynamic protein which supports cell
shape- defines cellular shape (without CS cell would not have polarity)
strength- imparts strength and prevents it from being damaged
movement- provides mechanism by which things can move within a cell and by which cells themselves can move

Question 2

cytoskeleton filaments

Answer 2

actin filaments- double stranded composed of actin protein subunits
regulate cell shape and locomotion
microtubules
intermediate filaments (IF)

Question 3

filaments

Answer 3

Actin filaments enable microvilli formation (important for nutrient absorption)

Actin also connects to cell junctions to maintain a physical barrier

Intermediate filaments (IF) anchor across the cell providing structure and strength via cell junctions called desmosomes

Microtubules run from basal to apical surfaces providing a intracellular transport network (polarity- have + and - end)

Question 4

cell polarity

Answer 4

refers to asymmetry; shape, structure, organisation
cell is not uniform

Question 5

intermediate filament structure

Answer 5

IFs held together by non covalent interactions eg h bonds, electrostatic
single protofilaments are thermally unstable as one break doesnt require much energy (very weak)

multiple protofilaments are thermally stable (strong)
breaking whole filament requires lots of energy to break bond

Question 6

actin structure

Answer 6

8nm diameter, thinnest filament
regulate cell shape and locomotion
F actin=filament G-actin= globular (monomer of which F actin is formed)
2 polymer chains form helix
can be found as linear bundles, 2D networks and 3D gels
has a large cleft in centre which binds ATP
G-actin subunits all point same way- polarity (have + end and - end)

Question 7

nucleation

Answer 7

first step in formation of a new structure via self assembly
2 ATP G actin subunits bind to each other weakly
when 3 ATP G actin subunits form a trimer it acts as a nucleus from which a large actin filament can form

Question 8

F actin kinetics

Answer 8

nucleation (lag phase)- actin subunits not yet in filaments
elongation (growth phase)- growing actin filament (once trimer has formed)
steady state (equilibrium phase)- actin filaments with subunits coming on and off

Cc- critical concentration (steady state)- G actin monomers and F actin are in equilibrium
rate of G actin addition and dissociation is equal at each end of actin chain
Gactin>Cc favours addition
Gactin<Cc favour dissociation

Cc is determined by 2 constants
Kon= rate of addition
Koff= rate of dissociation

Cc= Koff/Kon

Question 9

treadmilling

Answer 9

subunits are added faster at + end and dissociate faster at - end due to differences in Cc
causes + end to grow and - end to shrink
overall length of F actin remains constant - remains in equilibrium
essential for cell motility and alterations in cell shape

Question 10

actin motor proteins

Answer 10

F actin can form interactions with motor proteins eg myosin, kinesin, dynein
assists in movement

myosin binds to actin
ATP hydrolysation releases energy
mysosin pulls itself along F actin molecule
results in movement (muscle contraction)

myosin discovered in skeletal muscle, arranged in a way that myosin heads oppose each other and can pull actin filaments closer to each other
in skeletal muscle, treadmilling is prevented by binding proteins such as tropomudulin

Question 11

microtubules

Answer 11

long hollow cylinders of globular protein tubulin with central lumen
regulate cell transport
largest filament
each protofilament formed of repeated αβ tubulin heterodimers
13 longitdinally repeating units- long strands called protofilaments containing hollow lumen
helical structure
vertical interactions- α- and β-proteins
lateral interactions- α/ α and β/ β
microtubules very stiff
polarity- β-tubulin exposed at the (+) end
α-tubulin exposed at the (-) end

Question 12

microtubules formation

Answer 12

nucleation- within microtubule organising centres
MTOC called centrosome
(-) end remains anchored to centrosome + grows out of centrosome (+) end first

Question 13

centrosome

Answer 13

two features
-pair of centrioles
-pericentriolar material
organised structures consisting of 9 sets of triplet microtubules
orientated at 90 degrees to each other
- end anchored and + end pointing outwards

Question 14

petricentriolar material

Answer 14

amorphous material - disorgansies area of proteins
contains gamma tubulin ring complex ( gamma turc)
consists of multiple copies of y tubulin complexed with other proteins
y turc is specific site for nucleation for microtubules

Question 15

microtubule dynamics

Answer 15

have very short lifespan
polymerisatoon kinetics are similar to that of F actin
Cc is lower at + end and higher at - end so + end grows and - end shrinks (treadmilling)
dynamic instability- causes by premature hydrolysing of gtp in terminal heterodimer
as filament is growing, monomers have gtp, hydrolysis of gtp guarentees to add next monomer, if hydrolysis occurs before next monomer is added, causes shrinking back effects

Question 16

microtubule motor proteins

Answer 16

responsible for movement along microtubule
kinesin- moves towards +ve end of microtubules (anterograde- away from centre)
dynein- moves towards -ve end of microtubule (retrograde- towards centre)

Question 17

kinesin

Answer 17

starts with microtubule surface and kinesin molecule
lagging head is ATP bound- strongly associated with MT
leading head is ADP bound- weakly associated with MT
hydrolysis of ATP causes lagging head to dissociate from MT
leading head binds to ATP, causes conformational change (still bound to surface of MT)
lagging heads move forwards and binds further down MT
hand over hand mechanism
how cargo gets transported along MTs

(dyenin similar mechanism but move golgi vesicles)

Question 18

intermediate filament (IF)

Answer 18

provide mechanical strength (10nm thick)
multiple types composed of different subunits
much greater tensile strength than others
non polar- symmetrical
not associated motor proteins- not involved in cellular movement
great heterogeneity (lots of types) within IF

Question 19

hierarchical structure of IF

Answer 19

alpha helical region of monomer
forms coiled coil dimer
staggered tetramer of two coiled coil dimers
lateral association of 8 tetramers
addition of 8 tetramers to growing filament
provides lots of strength
eg in nuclear lamina

Question 20

tissues

Answer 20

multicellular organised structures
can withstand and respond to external cues
cells must link to eachother and to extracellular matrix (ECM)

categories- epithelial, muscle, nervous, connnective

Question 21

epithelial

Answer 21

sheets of cells- line surfaces of bod
ecm is present as a very thin sheet called basal lamina

Question 22

connective

Answer 22

these cells produce ECM and are sparsely populated within dense matrix

Question 23

types of cell junctions

Answer 23

anchoring junctions- anchor cytoskeleton between cells or between cells and ECM

occluding junctions -prevents passage of ions and small molecules between cells. typically tight junctions in vertebrates

communicating junctions- direct connections between cytoplasm of two cells- gap junctions

Question 24

anchoring junctions

Answer 24

depend on transmembrane (across membrane) adhesion proteins
-one end binds to cytoskeleton
-other ends binds to something extracellular

these cytoskeletal linked transmembrane proteins belong to two superfamilies
cadherins- cell cell attachment
integrins- cell matrix attachment

Question 25

cadherins

Answer 25

c terminus binds to cytoskeleton (filaments in cell)
n terminus binds to other cadherin
cadherins bind to same type of cadherin (homophillic adhesion)

all have extracellular portions containing multiple cadherin domains (EC domains)
almost all single pass transmembrane protein- only cross the membrane once
homophillic binding occurs at N terminal tips (last EC domain)

Question 26

actin binding

Answer 26

cadherins bind to cytoskeleton in cell
intracellular binding to cytoskleton is indirect and done via adaptor proteins- main type is catenin
assemble on C terminus (within cell) of cadherin and mediate binding
catenins link cadherin to F actin
link is called adherins junction

Question 27

adherens junctions

Answer 27

membrane protusions initiate cell cell contact
-small cadherin and catenin cluster
actin and cadherin recruitment expands junction
-additional cadherin and catenin
actin remodelling and myosin recruitment expands adherens junction
-contractile actin and myosin bundle - lined up near CM bringing cells closer together

adhesion belt
responsible for formation of tubular structures + blood vessels

Question 28

extracellular matrix

Answer 28

intricate network of proteins, carbohydrates and water (bone is exception)
provides support for cells and tissues
important regulator of cellular signalling (comms)
secreted by cells (fibroblasts)
organisation is tissue dependent, giving rise to specific tissue properties
eg bone- hard, tendons- strong, cornea- transparent

Question 29

ECM composition

Answer 29

glycosaminoglycans (GAGs)- oligosaccharides covalently bonded to a protein= proteoglycans
~36 types

fribrous proteins- predominantly collagens
~40 types

glcyoproteins- ~200 types

Question 30

cell matrix junctions

Answer 30

-focal adhesions (actin linked)
-hemidesmosomes

transmembrane cell adhesion proteins act as matrix receptors (proteins part of the integrin family)

Question 31

anchoring junctions- integrins

Answer 31

integrins composed of 2 glycoportein subunits (alpha and beta)
integrins bind actin (inside cell) to extracellular matrix
both subunits span the plasma membrane (single pass)
small intracellular C terminus, large extracellular N terminus

Question 32

integrin types: focal adhesions

Answer 32

focal adhesions in extracellular domain bind integrins to specific amino acid sequences in ECM proteins
fibronectin is most usual protein with RGD seqeunces (arginine, glycine, aspartate)
intracellulular domain binds to a complex of several proteins which link to F actin

Question 33

junction types: hemidesmosomes

Answer 33

junctions connect the ECM to IF within a cell
use integrins the same way as actin linked junctions
most prominent cell matrix junction in epithelial cells
rely on a specific integrin- α6β4

Question 34

intergin switching

Answer 34

switch between active and inactive so that cells are not always bound to extracellular matrix

inactive- N terminal domains are folded together so they cant bind matrix proteins + cytoplasmic tails are hooked preventing cytoskeletal binding

active- cytoplasmic tails unhook and expose them to adaptor proteins at n terminus- tails unfold and extend
strong ligand binding

Question 35

occluding junctions

Answer 35

known as tight junctions
seal the gap between epithelial cells
ensure that molecules/fluid entering at apical side cannot diffuse back peripherally- back and forth
formed of sealing strands - long rows of transmembrane homophillic adhesion proteins (cross memebrane back and forth)
tight junction proteins;
claudins- essential mediators of tight junction formation, key protein in pulling cells close together
occludins- non essential, limit permeability of junction

Question 36

communicating junctions

Answer 36

known as gap junctions
provide channels which connect cytoplasm of two cells
2 cell membranes come very close to one another and gap is spanned by channel forming proteins
connexins (vertebrates)
innexins (invertebrates)
protein assemblies called connexons penetrate adjacent cell membranes forming aqueous channel
6 connexins form a connexon
differences in connexin subunits regulate its permeability
permeable to small molecules eg ions, amino acids
macromolecules cant pass
sensitive to voltage, pH, neurotransmitters, Ca2+

Question 37

cadherin binding

Answer 37

occurs outside the cell
homophillic binding occurs at N terminus between Knobs and Pockets (ball and socket like)
cadherins have flacid (can’t bind) and erect states
ca2+ binds in hinge regions of cadherin between EC domains to keep cadherin rigid (allowing binding)
cadherins bind weakly (strand swapping) to eachother so pair up with other cadherins in parallel to form adherens junctios (multiple anchoring to plasma membrane)

Question 38

desmosomes

Answer 38

only found in vertebrates
found in tissues that are subject to mechanical stress eg heart and skin
link cells through intermediate filaments (IF)
linked to very strong filament which allow tissues to remain intact + prevent damage
same idea as linking to actin but stronger due to IF