Cell Biology of Disease Flashcards
Define dynamic instability
Rate of GTP hydrolysis
Determines alternation between shrinkage and growth of MTs
Rapidity of the two causes rapid turnover of most MTs
Give four diseases (tauopathies) caused by mutations in Tau
Alzheimers
Progressive supranuclear palsy
Cortico-basal degeneration
Frontotemperal dementia and Parkinson linked to chromosome 17
What are MAPs and what do they play a role in?
Stabilisation and destabilisation of MTs
(Needed for efficient cell trafficking)
Determining cell shape and polarity
When happens to Tau in Alzheimer’s disease
How can the NFTs be scanned?
Tau is hyperphosphorylated
No longer holds MTs together
Tau forms pairs helical fragments which develop into neurofibrillary tangles
Using positron emission tomography, visualisation of a chemical that binds to Tau
What is Lissencephaly caused by? What is the physiological consequence because of this?
Tubulin mutation Alpha-Tubulin, tuba-3 Results in tubulin misfolding which requires a series of chaperones Absence of normal folds in the cortex Otherwise called smooth brain
Give five symptoms of Lissencephaly
What is life expectancy?
Abnormally small head Unusual facial appearance Difficulty swallowing Failure to thrive (muscle spasms, seizures, severe psychomotor retardation) Most die before age of 12
What was the first discovered mutation for Lissencephaly?
Missense
Arg264 to Cys
For Lissencephaly dimers of tubulin can form and polymerise normally
What could be the cause of the disease instead
Incorrect binding to other MT binding proteins
What are MTs made of?
How big are they
Alpha and beta tubulin dimers
25nm diameter tubes
How many kinesin genes are there in humans?
40
What is hereditary spastic paraplegia? What mutation is it caused by
Kif5a, kinesin-1
Mutations in motor domain
Describe symptoms of hereditary spastic paraplegia
What kind of disease is it?
Progressive weakness and stiffness in legs
Average age of onset 24 years
Rare disease
Autosomal dominant
Missense mutations, single amino acid change
Name different kinds of mutations and what they cause
N256S and K253N
Reduced velocity of movement
R280S and K253N
Reduced binding to MTs when attached to cargo
A361V
No change in vitro
What drug inhibits mitotic kinesins?
Which kinesins does it inhibit
Monastery
Eg5
Briefly describe actin
Globular monomer that polymerises into filaments 375 amino acid Molecular weight 42 kDa Three main types Alpha in muscle Beta and gamma in non muscle cells
Describe structure of F actin
Two long pitch helices
One pitch every 5.5nm
72nm long
Where and how are actin and titis molecules joined in muscle?
Cross-linked in the Z-disc by a Z line protein alpha actinin
What is the function of Titin?
Provides binding sites for numerous proteins
Acts as a molecular spring for passive elasticity of muscle
Give two examples of proteins with mutations that may cause be alone myopathies
What do these cause lack of?
Actin and nebulin
Lack of sarcomeres meaning muscle is weak
Death from effects on respiratory muscle
Symptoms of nemaline myopathy
Facial weakness Scoliosis Floppy babies Can't sit or stand May need ventilation Respiratory problems Multiple joint contractures
What do these mutations cause in terms of types of congenital myopathies? G15 D154N V163L I136M D292V
Actin myopathy — severe weakness, high mortality
Actin myopathy — some nemaline and IRM, severe, high mortality
Nemaline and IRM — less severe
Nemaline and IRM — less severe
Congenital fibre type disproportion
Describe duchenne muscular dystrophy
X chromosome gene X- linked recessive Mutations in dystrophin Most affect boys, 100 per year in UK wheelchair bound by 10, death from cardiac problems by 20
What is dystrophin?
Binds gamma-actin and alpha-beta dystroglycan
Found in muscle costameres (close to plasma membrane in skeletal/cardiac muscle)
What is the function of dystrophin?
What do mutations in it cause?
Links internal cytoskeleton to extra cellular matrix
Dystrophin loss causes muscle wasting
Affects connectivity between z-discs
Muscle less able to withstand damage upon contraction = membrane ruptures
What other mutations in actin based cytoskeleton cause diseases and what diseases do they cause?
Myosin 2a - blood disorders in platelets/WBC
Myosin 7a - deafness and blindness (usher syndrome)
Myosin 5a (griscelli syndrome)
Briefly describe intermediate filaments
Rope like 10nm diameter Tough and durable Form nuclear lamina, also in cytoplasm Not very dynamic, Central rod domain important
Name four types of intermediate filaments and their cell location
Keratin - epithelial
Desmin - muscle
Vimentin - fibroblasts
Neuronal intermediate filaments - neurones
What are desmosomes?
Localised spot-like adhesions randomly arranged on lateral surface of plasma membrane
Use desmosome in
Help resist shearing forces
What are hemidesmosomes?
Also called focal adhesions
Where cells attach to underlying extracellular matrix
Connect basal cell face to basal lamina
Use desmopenetrin
Mutations in desmin affect skeletal, cardiac and smooth muscle
Name and briefly describe two main heart problems from this
Hypertrophic cardiomyopathy - thickened portion of myocardium causes sudden cardiac death
Dilated cardiomyopathy - portion of myocardium dilated
Heart becomes weakened and enlarged, less blood pumping efficiency
Which genes cause nuclear envelop aphids (lamin loathes)
Lamin A and C
What are the roles of nuclear lamin?
Scaffold for protein complexes
Regulate nuclear structure and function
Give an example of a Lamin A mutation
E145K in lamin A central rod
Affects filament formation/polymerisation
Causes down regulation of B lamin
Causes premature senescence
How are proteins recognised for translocation into the ER?
N terminal signal sequence
Short and hydrophobic
Cleaved by signal peptidase
Retained for transmembrane proteins
What is the Sec61 translocon?
Forms aqueous pore in ER membrane upon displacement of SRP
translation occurs simultaneously with translocation into the ER
What function does the oxidising environment of the ER have for protein folding?
Promotes the binding of cysteine residue and formation of disulphide bonds
Promoted by protein disulphide isomerise
The cytosol is reducing enviromemt
What does ER glycosylation entail?
Addition of pre formed oligosaccharide to asparagine residues
2N-acetyl glucosamines, 9 mannoses 3 terminal glucoses
Describe protein folding after glycosylation
Trimmed leaving 1 glucose
Chaperone calnexin and ERp57 bind (protein and cysteines) preventing and aggregation
Release of protein removes final glucose
Incorrect folding adds glucose using glycosyl transferase for recognition by Valencia
How are unwanted proteins recognised for ERAD
Mannose residues are trimmed
Retro-translocated through sec61 and poly ubiquitinated
Then degraded by cytosolic proteome complex
How are escaped membrane protein returned to the ER?
Contain KKXX motif recognised by interaction with coatamer complex - COP1
Transport vesicles return to ER
How are escaped luminal proteins returned to the ER?
Contain KDEl motif
Recognised by coatamer and returned in COP1 vesicles
What is Cf caused by? How many affected/carriers?
What is the most common mutation for CF?
Loss of function of cystic fibrosis transmembrane conductance regulator
Normal function as an ATP binding cassette, couples ATO hydrolysis with Cl- transport
1/27 carry mutation, 1 in 2000 affected
Autosomal recessive
3nt deletion loss of phenylalanine
DeltaF508
At least one mutated in 90% of cases
Name and briefly describe 2 other diseases associated with protein folding
Osteogenesis imperfecta
Collagen I mutations prevent folding, cause degradation of pro collagen
Severe defects in bone formation
Alpha1-anti trypsin deficiency
Mutation Impairs folding
Cause protease damage in lungs resulting in emphysema
How does the cholera toxin enter the ER starting from outside the cell?
Promotes electrolyte/water movement into intestinal lumen causing diarrhoea
B subunits binds to GM1 on gut surface
Alpha subunit possesses KDEl motif, enters the ER
Dissociates into 2 subunits
Retro translocated avoiding degradation, too few lysines
Binds adenylate cyclase, increases cAMP which activates protein kinase A
PKA Phosphorylated CFTR causing CL- transport
How does ricin toxin get to the ER from outside the cell and what symptoms does it cause?
Symptoms = nausea! diarrhoea! seizures and hypotension
Same as ER
B subunit binds via b-galactose residues on glycol iodide
Retrograde transport to ER
A subunit possesses KDEl motif,
A and B dissociate, A unfold and retro translocated across membrane avoiding degradation by too few lysines
Target 28S rRNA and depurinates it by cleavingA4324 and releases adenine
Site of elongation factor binding - inhibits protein translation
How does HMCV US2 and US11 avoid detection by the immune system
US2 and 11 are localised to the ERr membrane
They mark MHC1 molecules for ERAD pathway
No MHC1 molecules can then be produced to detect the virus
How does adenovirus affect MHCI molecules
Adenovirus E3/19K bind to MHC1
Protein has cytoplasmic tail KKXX motif for retrieval of MHC1 at cis Golgi
Returns to the ER and prevents it from reaching the plasma membrane
Give five characteristics of lysosomes
All mammalian cells except RBCS About 1% of cell volume Roughly spherical Dense protein rich core 200-400nm diameter
Lysosomes are involved in…
Degradation of molecules by endocyotosis or auto phage
Apoptosis
Plasma membrane repair
Secretory organelle in immune cells
How many hydrolysis do lysosomes contain?
Normal mode of degradation?
Over 45
Covalent bond hydrolysis
(Nucleases, proteases, glycosidases, lipases)
What is the pH of lysosomes and how is it maintained?
pH of cytosol?
PH between 4.5 and 5.0 maintained by action of H+ vacuolar pump powered by ATP hydrolysis
Cytosol pH 7.2
How do lysosomes pump degradation products into the cytosol?
Using secondary active transporters coupled to proton pump coupling
Transport water soluble molecules = sugars! amino acids and nucleosides
Reuse for macromolecules
Material for lysosomal degradation is transported in which three ways?
Endocytosis (fluid phase mols, plasma membrane proteins)
Phagocytosis (large extracellular species such as microorganisms)
Autophagy (cytosolic, whole organelles)
How do molecules not meant for degradation get back to the plasma membrane
Recognised by the transferrin receptor
Lysosomal degradation plays an important role the down regulation of receptor signalling
What is the best characterised form of autophagy?
What does it involve?
Macro autophagy
Envelope donated from plasma membrane or organelle surrounds cytoplasmic material forming an auto phagosome which binds with lysosomes
Which drug can be used to enhance macro autophagy?
What disease does it show uses in?
Promoted by starvation of the cell achieved by inhibition of the regulatory protein, the mammalian target of rapamycin/mTOR
Rapamycin promotes clearance of Hungtingtin aggregates in mice and some mutant forms of alpha-synuclein in Parkinson’s disease
How are lysosomes involved in apoptosis
Apoptosis is indeed by binding to cellular targets of proteases called caspases
Apoptosis cells observe increase permeability of lysosomes membrane
This releases lysosomes proteases called cathepsins that also act on on cellular targets
Can work at cytosolic pH, and trigger mitochondrial intrinsic pathway apoptosis via Bid
How are lysosomes involved in plasma membrane repair?
Plasma membrane can be damaged by mechanic damage
Lysosomes help to repair these holes
Damage releases Ca2+ detected by synaptotagmin 7 on lysosomal membrane
In which 3 ways are lysosomal molecule accumulation problems associated with diseases?
How many genetic diseases are associated with this?
Insufficient hydrolyse transport
Deficiencies in lysosomes
Inability to deliver breakdown products
Over 50 genetic diseases
What are the symptoms of I cell disease
What is the molecular basis I.e. What does the mutation do?
Think in terms of the 3 lysosomes genetic diseases
Also called mucolipidosis type II
Autosomal recessive
Mutation in GNPTAS
N-actylglucosaminidase-1-phosphotransferase
Causes formation of inclusion bodies
Symptoms = facial/skeletal abnormalities, psychomotor retardation, heart failure in first decade
Hydrolyse monosylated Glycans are not modified by the enzyme and so are not transported to endosomes and then lysosomes but instead into the cytosol
What are the symptoms of Pompe disease?
Describe what effect the mutation has on lysosomes and its STORAGE
Autosomal recessive
Hydrolase alpha-d-glucosidase
Symptoms = progressive cardiac/skeletal myopathy, cardio respiratory failure within 1st year
Small %of cell glycogen enters lysosomes for breakdown into glucose
No working enzyme = glycogen accumulation in lysosomes
Enzyme replacement only works in cardiac tissue, taken up by mannose-phosphate receptors
What are the clinical symptoms of Fabry disease? What kind of mutation does it cause?
What does it result in the accumulation of?
X-linked disorder, alpha-galactosidase
Symptoms = facial abnormalities, non specific effects for renal/cardiac problems, progressive organ/tissue damage
Renal treatment increases life from 40 to 50
Enzymes usually removes terminal galactose from GB3
Globotriaosylceramide
Accumulates in kidney tubules and glomerular cells, nerves and dorsal root ganglia
Enzyme replacement
Describe infantile Sialic acid storage disease and Salla disease
The symptoms and molecular basis
Autosomal recessive mutations in Sialin gene
Symptoms = facial abnormalities enlarged heart/liver/spleen, mental retardation and death within first 2 years
Salla = Finnish population! less severe! life expectancy 50
Normal transport of Sialic acid (9carbon monos) that are breakdown products of glycoproteins/lipids/saminoglycans
Abnormal accumulation in lysosomes
Briefly describe the nucleus
Function, four seperate structures
Contains genetic material
Maintain integrity of genes, control cell activities by regulating gene expression and replication mediation
6 um, 10% cell volume
1) envelope
2) membrane, impermeable to large molecules
3) nucleoskeleton/lamina, meshwork adding mechanical support
4) subnuclear bodies,
Briefly describe the nuclear envelope/membrane
Double lipid bilayer
Outer membrane contiguous with rough ER
Inner membrane connected to lamina and intermediate filament network
Peri nuclear space is 20-40nm
How thick is the nuclear lamina and what is its function?
Dense fibrillation network, 30-100nm
Mechanical support, regulate replication and cell division, chromatin organisation, anchors nuclear pore complexes
What are laminopathies? What mutations cause?
Give but don’t describe two examples
Mutations in lamin A and C and associated proteins e.g. Emerin
Defects in filament assembly and/or attachment to nuclear envelope, jeopardises nuclear stability
No cure
What is Emery-Dreyfus muscular dystrophy? What kind of disease? What are the symptoms?
Affects skeletal/cardiac muscles
Causes joint deformities called contractures restricting joint movements
Slowly progressive muscle weakness/wasting
Abnormalities in heart electric signals, abnormal hearth rhythms
What kind of disease of Hutchinson-Gilford progeria syndrome? What mutant protein is its cause?
Physical aspects of ageing accelerated
Point mutation in LMNA gene, LAD50 mutant protein incorporates abnormally into lamina (lacks 50 amino acids)
Mechanical defects, lamina thickening, loss of peripheral heterochromatin, increased DNA damage
