Cytoskeleton Flashcards
actin filaments, intermediate filaments and microtubules
Main functions of cytoskeleton (8)
- provides cell shape through the maintenance of asymmetry
- chromosome separation during mitosis
- vesicle trafficking and organelle positioning
- supports plasma membrane to bear stress
- cell motility/migration
- muscle contraction
- movement of cilia and flagella
- maintenance of axons and dendrites in neurons
3 types of filaments of the cytoskeleton and their sizes
- actin microfilaments (7nm)
- intermediate filaments (10nm)
- microtubules (25nm)
how can the cytoskeleton be visualised?
antibodies and fluorescence –> this can test for pathologies because a defect in the cytoskeleton would alter the morphology of it
dynamic vs stationary filaments definition
DYNAMIC: the length of the filaments is continuously modified (lengthening and shortening)
STATIONARY: do not possess dynamic instability and are used to support and strengthen the cell junction
which filaments are dynamic and which are stationary (or both)
ACTIN: S and D
INTERMEDIATE: only S
MTs: S and D
G vs F actin definition
G -actin: globular actin that is free in the cytoplasm
F-actin: fibrous actin that is polymerised on the filament
Structture of actin filament
-long chain of G-actin monomers connected in a ‘string of pearls’
-2 ends:
1. positive (barbed) end which is where addition of monomers occurs
- negative (pointed) end where there is release of monomers
how is the length of actin filaments controlled?
using the rate of monomer polymerisation at +ve end vs monomer release at -ve end:
- addition> release: lengthens
- release>addition: shorterns
- addition = release: constant length known as actin treadmilling
process of actin monomer polymerisation
-occurs at positive end
-requires K+, Mg2+ and ATP
-ATP hydrolysis occurs when the monomer is added but the Pi group is not released immediately and this forms a transient section of ADP-Pi bound actin that is detectable until the release of the Pi
differences in actin found in different cells
6 actin genes = 6 isoforms: 2 are ubiquitously expressed
alpha/gamma smooth actin, alpha skeletal and alpha cardiac
-epithelial cells have actin as microvilli (apical specialisations) arranged in parallel bundles
-present in sarcomeres
-pseudopods and lamellipodia
-dynamic contractile rings
!!! actin is only stationary in microvilli and sarcomeres
How were the two ends of actin defined (experiment)
-actin was allowed to bind with HMM (heavy meromyosin containing the globular heads)
-formed an arrowhead structure where the HMM was bound at 45 degree angles
-the pointed end of the HMM was found at the -ve end and the open part of the arrow (called barbed) was found at the +ve end
Filament bundling proteins function
cross linkage of fibrils to form parallel bundles to provide support and integrity
eg: fimbrin and fascin
General types actin binding proteins (5)
- cross linking
- filament severing
- motor
- capping
- filament bundling
Filament severing proteins function
proteins that cut long actin filaments into smaller fragments
eg:
1. gelsolin: at high concs of Ca2+ ions they act as capping proteins
2. cofilin: severs filaments into free positive and negative fragments
actin Capping proteins function
bind on ends of actin molecules to regulate the length either by preventing polymerisation (short) or by preventing monomer release (long)
eg:
tropomodulin: caps the negative end to prevent dissociation of monomers
cappZ: caps positive end to prevent monomer association
Formine: blocks the binding of proteins interactive with the positive end so it can grow (no inhibition of polymerisation)
Filament crosslinking proteins function
cross linkage of actin filaments with eachother
eg:
spectrin, adducin, protein 4.1/4.9
Actin-motor proteins function
belong to the myosin family and allow the hydrolysis of ATP so energy is provided for the movement of actin monomers through the length of the filament
organisation of actin filaments in epithelial lining cells
-contain cortical actin (and actin if they have microvilli)
-terminal web: region beneath plasma membrane where the actin is thicker (this allows interaction with intermediate filaments)
main functions of actin filaments
- cell motility and locomotion
- anchorage and movement of membrane proteins
- microvilli core and presence in the terminal web of epithelia
- extension of cell processes (formation of podia)
How does actin allow cell locomotion?
ABM: ACTIN BASED MOTILITY
- achieved by the force exerted by actin filaments undergoing polymerization at their growing ends
leading edge: cells extend processes by pushing the plasma membrane ahead of the growing actin filaments
trailing edge: depolymerisation of actin causes its retraction
CYCLE OF MOVEMENT:
-elongation of membrane
-new adhesion to surface
-translocation in direction of movement
-de-adhesion/retraction of the adhesion furthest away from the direction of movement
Main properties of intermediate filaments (4)
-non polarised
-stable and strong (strongest out of the three cytoskeletal components)
-only stationary (no dynamic)
-composed of different proteins in different cells
general functions of intermediate filaments
-provide mechanical stress to cytoplasm
-help epithelial layers resist shear stress
-present in cell junctions (desmosomes and hemidesmosomes)
-hold the nucleus in its required position
detailed structure of intermediate filaments (up to tetramer stage)
-monomers are helical in nature and are composed of a globular head on each side - one COOH and one NH2
-coiled dimers are formed by the twisting of two monomers so that the NH2 and COOH heads are associated with their like counterparts
-tetramers are formed by the antiparallel coiling of the dimers so that the NH2 and COOH regions are now facing eachother. This eliminates polarity because the opposite charges cancel out.
These tetramers are staggered throughout the rope like molecule
intermediate filament proteins found in different cell types (NEED MEMORISATION) –> (6)
- epithelial cells: basic and acidic cytokeratins
- mesenchymal cells: vimentin
- muscle cells: desmin
- neuroglial cells: GFAP (glial fibrillary acidic protein)
- neurons: neurofilament proteins
- nucleus of all cells: lamins (A/B/C)
Pathology associated with cytokeratins
EPIDERMOLYSIS BULLOSA SIMPLEX
characterised by the formation of blisters induced by even small amounts of mechanical stress, because there is no keratin present in cell junctions and this allows the cells to break apart
Keratin proteins details
-present in epithelia
-connect desmosome and hemidesmosome junctions
-mass produced by the differentiation of the epidermis when producing the stratum corneum layer (outer skin protection)
-found in skin, hair and nails
Desmin proteins details
-present in muscle cells
-prevents overstretching
-aligns the Z lines of the sarcomere, hence holding it in place
-if it is dysfunctional we observe a deranged myofibril architecture
how can we use intermediate filaments in tumour diagnosis?
-intermediate filaments are highly tissue specific and so to determine the origin of a tumour we observe the characterisation of intermediate filaments within the cell cytoplasm
- CARCINOMA: epithelial tumours that are characterised by presence of keratins
- SARCOMAS: connective tissue tumours that are characterised by the presence of vimentin (or sometimes desmins)
General functions of microtubules (5)
- creation of system of connections to allow intracellular vesicle transport
- cilia and flagella motility
- attachment of chromosomes to the mitotic spindles
- maintenance of cell shape (and asymmetry)
- regulatory effect on cell elongation and movement
structure of a microtubule
-hollow cylindrical tube
-composed of 13 protofilaments (strands of proteins) arranged in a circle
-each protofilament is made of dimers of connected proteins: alpha and beta tubulins
-the negative end of MT contains gamma tubulin (needed for MT nucleation), capping proteins and the MTOC
-25 nm in diameter
-polarised and can be either dynamic or stationary
what factor determines the dynamic instability of MTs (2)
- abundance of tubulin substrates in environment
- MAP proteins
Process of MT polymerisation
-monomers are added at the positive end and are released in the negative end
-tubulin bound GTP favours polymerisation and tubulin bound GDP favours depolymerisation
-controlled by MAPs
Role of MAP proteins in the regulation of MT stabilisation
MAP BOUND TO MT: blocks dynamic instability and allows them to be more stable and longer
MAP UNBOUND TO MT: dynamic instability becomes more evident so MTs become shorter and more dynamic
what type of equilibrium is reached in MT dynamic stability?
the overall concentration of MTs stays constant, however their length constantly changes (lengthening and shortening of MT sometimes to the point where MTs disappear completely)
structure of MTOC
MTOC: microtubule organising center
-region of the cell where MTs are formed
-contains hundreds of rings of gamma tubulin and 2 centrioles
location of MTOC
!! correlation between MTOC location and cell morphology:
- located in nucleus in non dividing cells
- located in poles of mitotic spindle in dividing cells
- located in cilia attached to basal bodies
- located in the space between nucleus and cell body in neuronal axons
Structure of the centrosome
1 centrosome = 2 centrioles (one mother and one daughter):
-orthogonally arranged
-surrounded by amorphous tissue containing pericentriolar material and gamma tubulin
-nucleation site: the negative end of the microtubule which is embedded into the centrosome
Structure of the centrioles
-hollow cylinder
-wall made up of 9 triplets of MTs (A,B,C)
- A MT is a complete MT made of 13 protofilaments
-B and C MTs are non complete MTs made of 10/11 protofilaments and hence need to be connected to eachother + the A MT to ensure their completion
MOTHER CENTRIOLE: completely hollow, contains external distal and sub-distal appendages
DAUGHTER CENTRIOLE: not completely hollow, as it contains a protein made cartwheel structure in its lumen which appears electron dense in an EM
microtubule catastrophe def
the term used to describe the process of suddenly switching from a growing to a shortening MT
Microtubule motor proteins
-attach organelles to the MT tracks in cells
-energy is required (ATP)
2 MMP families:
1. DYNEINS: move towards the negative end of MT
-move organelles from the cell periphery to the MTOC
- KINESINS: move towards positive end of MT
-move organelles from the cell center to the cell periphery
! both also play a role in mitosis/meiosis
What are NBBCs?
nucleus basal body connectors –> link centriole to the mitotic spindle poles during mitosis
Describe the stages of the centrosome cycle (7)
- disengagement: tight link between mother and daughter is severed and they are only held by loose fibrous connections
- duplication: during S PHASE
- engagement: new centrioles formed reach full length, daughter becomes mother
- maturation: collection of pericentriolar material
- separation: during PROPHASE, generation of spindle fibres and centrioles move away from eachother
- bipolar spindle formation: METAPHASE, centrioles at opposite poles of cell
- cell division: each daughter cell receives one centrosome during division (ie, a pair of centrioles)
DDE, MSFD
CILIA function
-apical specialisation of lining epithelium cells
-contain MTs
-have the property to bend and move materials located on the apical surface of cells
general regions of MTs in cillia (most apical to most basal)
- expanded tract: free portion that is able to bend
- transitional zone containing basal plates: allows implantation to the apical cytoplasm
- basal bodies: centrioles whichanchor the axoneme
- protein rootlets: anchor the Ts deeper into the cytoplasm
structure of the axoneme
-9 doublets of peripheral MTs, one having a full 13 protofilament shell and the other having 11 and being completed by the first one
-2 central MTs surrounded by a central sheath
-MTs linked by:
1. dynein arms (MAP)
2. nexin linking proteins (connects peripheral MTs with eachother)
3. radial spokes (connects peripheral to central MTs)
Process of cilia movement
-mediated by the dynein arm proteins (MAPs)
-uses Mg2+ and ATP to pull on cilia and bend them
pathologies associated with faulty cilia
PRIMARY CILIARY DYSKINESIA: dynein arms do not properly work and cilia doesn’t bend properly
-affects dust removal in airways
-affects spermatozoa motility
