Lecture 4 - The cytoskeleton (part 2) Flashcards
What are two key intermediate filaments and what roles do they play in the cell#?
-keratin
makes up the cytosolic intermediate filaments that extend from the nuclear membrane to the cell membrane e.g. in skin epithelial cells
-laminin
makes up nuclear intermediate filaments
What are key features of intermediate filaments?
- divergent in sequence and size (unline microfilament and microtubules)
- large diversity (unlike mf and mt)
- 5 major classes based on sequence similarities, includes hard keratins from skin and nails
What are features of the keratins, and what type are they?
Intermediate Type I (acidic keratins) and Type II (basic keratins) filaments
- in epithelia
- heterodimers of basic and acidic subunits
- prominent in skin hair and nails
- mutations are involved in skin disease
What are the features of Vimentin and what type of intermediate filament are they?
Type III
-widely distributed (lymphocytes, endothelial cells and fibroblasts)
-role in structure to support the cell membranes and keep nucleus and organelles in place
-
What are the features of Neuronal IF proteins and what type of intermediate filament are they?
Type IV
- Found in neuronal neurofilaments
- determine the axon diameter and therefore the speed of conductance
- have a structural role in axons
What are the features of laminins and what type of intermediate filament are they?
Type V
- found in the nucleus
- fibrous network
- role in regulating and supporting nuclear shape and organisation
- role in organising and packaging DNA
What is the process of intermediate filament assembly?
1- Monomer forms a hetero or homo dimer with another monomer, forming a parallel dimer with C and N terminal ends
2- These are stacked in an antiparallel arrangement to form an antiparallel tetramer
3- Antiparallel tetramers are stacked end on end to form an elongated filamentous structure = protofilament
4- 2 protofilaments come together to form a protofibril
5- Four protofibrils wrap around each other to form an intermediate filament
What is the structure of an intermediate filament monomer?
- globular NH2 head
- globular COOH tail
- 48nm
- Central alpha helical rod domain
- spacers in the alpha helical rod region between (non helical)
What are the features of intermediate filament assembly?
- does not requite ATP or GTP hydrolysis
- spontaneous
- coil domains wrap around each other to form a coiled coil
- globular domains project away from the intermediate filament
What is the structure of the epidermis and the keratins involved in the cells of the epidermis?
outer epithelial layer Stratum corneum layer stratum granulosum layer stratum spinosum layer statum basale (basement membrane) dermis
cells in stratum basale layer express K5/14 heterodimers, expressed less as cells leave layer. Begin to express K1 and K10 as they leave reflects changing activity
Give an example of a disease associated with mutations in keratins
Epidermolysis Bullosa Simplex (EBS) - Blistering disease
- autosomal dominant muation
- caused by mutations in K5/14, aggregates of keratin found in the cytoplasm
- epiidermis no longer attached to the basement membrane and dermis
- N- or C- terminal muations means cells are unable to form protofilaments and the entire intermediate filament cannot form completely
- cells at the base of the epidermis are weakened
- epidermis and dermis easily separated
Give an example of a disease associated with Laminin
Hutchinson-Gilford progeria syndrome
- rare premature again disease
- caused by mutations in the LMNA gene (150bp deletion) produces an abnormal form of the nuclear intermediate filament laminin A
- nuclear envelope becomes unstable and prone to damage
- > bone disease, hair loss, cardiovascular disorders, diabetes and muscle atrophy
What are microtubules and how are they arranged?
- polymers of globulun tubular dimers
- arranged into tube ~24nm diameter,
- role in transport - cellular tracks for transporting vesicles, organelles and chromosomes
- used by microtubule motor proteins, kinesin and dynein
What are microtubules composed of?
Alpha-tubulin and Beta-tubulin subunits
Alpha-tubulin binds GTP irreversibley can cannot be hydrolysed (non-exchangable GTP)
Beta-tubulin binds GTP reversibly and can be hydrolysed to GDP (exchangable GTP, done as tubulin grows)
What is Taxotene and what does it do?
Anti-cancer drug which targets the B tubulin found in microtubules to allow crystallisation of the tubulin dimer
What is the process of assembly of microtubules?
- Tubulin dimers assemble longitudinally into short protofilaments
- The protofilaments associate laterally into a curved sheet, increasing its stability
- Tubulin dimers continue to associate with the tubular structure
- Beta tubulin associates with the (+) end
- Alpha tubulin assocaites with the (-) - As the dimers bind, the GTP on B tubulin is hydrolysed, creating a GDP microtubule
- If the rate of addition is greater than the GTP hydrolysis then a GTP cap is produced. As the rate of assembly is faster at the (+) end
What is a microtubule GTP cap?
an area at the (+) end of a microtubule where GTP is maintained by alpha and beta tubulin
provides stability to growing structure and regulates length
What are some similarities between microtubules and microfilaments?
- dynamic interaction dependent on the Cc of the subunits within a cell
- GTP cap/capping proteins
What is the rate of microtubule growth dependent on?
the availability of dimers in the cell, the Cc of alpha and beta tubulin
- once Cc is reached tubulin dimers can be converted into microtubules
- if tubulin dimers are more than the critical concentration there is preferential addition of tubulin at the (+) end
- if tubulin dimers are less than the Cc there is preferential loss of tubulin at the (+) end
How are microtubules stabilised?
-GTP cap
How is microtubule stability regulated?
By 'dynamic instability' series of 'catashtopihe and 'rescue' events -Catastrophe - loss, quicker -Rescue regrowth Dependent on cell Cc of tubulin dimers
What is the structure of the centrosome?
2 centrioles
- organised at right angles
- made up of 9 sets of triplet microtubules
- surrounded by a percentriolar matrix = acts as a nucleation point and contains, gamma tubulin and pericentrin, centroles interact with the PC
What is the MTOC?
the microtubule organising centre -structure used to organise tubules in the cell -located near the nucleus -directs assembly and orientation of microtubules -directs vesicle traffic -directs orientation of organelles Animals = centrosome Plant = nuclear membrane, no MTOC
What is the process of nucleation by microtubules at MTOCs?
gamma tubulin rings acts as nucleation points
growth from the (-) to the (+) end
(+) end can be capped for stability
Where can the MTOC be found?
- near nucleus
- at the base of cell extensions e.g. cillia, flagella, dendrites (basal body, acts as an MTOC, structurally similar)
What are the importances of MTOCs?
Cell division
- allow spindle apparatus to form and organise microtubules and for the division of chromosomes
- In nerve cell, long extensions of dendrites have microtubules with mixed polarity for transport
What are the types of micrtubule motor proteins and what is their mode of transport?
Kinesins (Anterograde transport) -membrane vesicles out -mitochondrion in Dyneins (Retrograde transport) -lysosomes for degradation -organelles in for degradation
Can be regions in the cells with little of either microtubules or microfilaments and so single vesicles may have both kinesins and myosin motors attached
What is the structure of kinesins?
- dimer
- two globular head with ATP binding site for binding to microtubules
- coiled alpha helix
- tail end contains two light chains which bind to the transported vesicle
What are the similarites between kinesins and myosin?
- both have ATP binding regions
- both have interacting regions (Actin binding site/Microtubule binding site)
- head, neck and tail region
- made up of heavy and light chains
