13Lyso - Inherited Lysosomal Disorders Flashcards
What are lysosomes?
Lysosomes are specialised sites of intracellular digestion
They are produced in the Golgi
They acquire hydrolytic enzymes - which are specially trafficked to lysosome using mannose 6-phosphate tags.
Exocytosis is essential to their function.
When were lysosomes discovered?
Lysosomes were discovered by Duve in 1955.
What can lysosomes be described as?
They can be described as:
Cytoplasmic organelle of variable appearance
Single membrane bound
Interior pH low - pH= 4-5
Interior is topological equivalent of cell’s exterior
In what types of cells are lysosomes found?
Present in most cells, they bud off from the Golgi
What do lysosomes contain?
Contain exo-hydrolases = each target specific linkages = act sequentially = act on the ends of macromolecules
Combined action therefore allows for digestion of many unrelated macromolecules
What types of material do lysosomes digest?
Digest both exogenous and endogenous material
What is heterophagy?
HETEROPHAGY
general cell feature; endocytosis, pinocytosis and phagocytosis
rate can be extremely high in some cell types
What is autophagy?
AUTOPHAGY
general cell feature; target is usually a cellular organelle = process is used for degrading damaged cell organelles
What is crinophagy?
CRINOPHAGY
secretory cells only; secretory cells fuse with a primary lysosome
What are the membranous bodies formed by lysosomes called?
Digestive vacuoles
What is the result of heterophagy/autophagy/crinophagy by the lysosome?
Result:
either soluble monomers (sugars, amino acids etc) diffuse out via transporters
or insoluble/indigestible material disgorged into extracellular fluid or remains in secondary lysosomes
What are the general features of inherited lysosomal disorders?
“An inherited pathological condition inherited by a primary defect in one lysosomal protein” - Hers, H. G. (1965)
Many (>40) now known
General features include:
failure in intracellular vacuolar digestion
large amount of macromolecules; lack of single hydrolase can affect many unrelated molecules
material is intravacuolar
results in MECHANICAL interference with cell function - enlarged lysosomes
lack of enzyme seen in many tissues/cells = pleiotropic
progressive diseases
amenable to therapy (?) - heterophagy
recessive inheritance
How is N-glycan degradation affected in inherited lysosomal disorders?
Defects in specific enzymes can affect degradation
Many proteins N-glycosylated = require specific enzymes for turnover to occur
α-Neuraminidase
β-Galactosidase
β-Hexosaminidase etc.
What are GM2 gangliosidoses/sphingolipidoses?
Disorders in which sphingolipidoses cannot be turned over.
How do GM2 gangliosidoses/sphingolipidoses occur?
Gangliosides found in the OUTER leaflets of all animal cell plasma membranes
Highest concentrations found in neuronal plasma membranes = symptoms often neurological
Gangliosides degraded in lysosomes
First degradative step requires hexosaminidase A
If hexosaminidase A activity is lost, cannot turnover sphingolipids
Activator involved
Hex A = αβ heterodimer
Activator protein forms complex with substrate = makes substrate more accessible to the enzyme - by solubilizing it
Hex A is the only Hex which acts on GM2 gangliosides = need Hex A to degrade GM2 gangliosides
What are the common features of gangliosidoses?
All recessive
Three variants:
1) α subunit = Tay-Sachs Disease
No active Hex A (or Hex S)
Mutation heterogenous - mainly α0
α-mRNA absent = RNA instability
α-DNA present
Causes GM2 accumulation in neuronal lysosomes
Heterozygote frequency is around 1 in 27 in Ashkenazi Jewish population (around 1 in 300 in the general population)
2) Activator protein / AB variant
no degradation of GM2 although Hex A and B are both active
symptoms very similar to Tay-Sachs
very rare disorder = much rarer than Tay-Sachs
3) β subunit - Sandhoff Disease
No active Hex A (or Hex B)
Mutations heterogenous; both β0 and β+ exist
Causes GM2 and soluble oligosaccharides to accumulate in lysosomes
Heterozygote frequency is around 1 in 300
What are the common features of Tay-Sachs disease?
Children initially appear normal but then develop the following symptoms:
Developmental retardation (missed developmental milestones)
Paralysis
Dementia
Blindness
Typically fatal in 2nd or 3rd year of life
Clinically: presents with a characteristic “cherry-red” spot on the retina = build up of GM2 sphingolipids
Pathologically: characteristic “ballooned” neurons in the brain and signs of spongiform encephalopathy = large histological changes
Concentric rings in lysosomes can be observed = formed by sphingolipids forming membranes = occluding lysosomal space
What is the origin of Tay-Sachs disease?
Arisen out of parts of Eastern Europe which had a higher TB load than other areas
Heterozygosity may provide some protection against TB infection
What is Gaucher’s disease?
First observed in 1882 = one of the oldest lysosomal storage disorders indentified
Accumulated material is glucosylceramide.
Defect identified as mutation in glucosylceramide β-glucosidase (glucocerebrosidase)
Gene located at 1q21
Neighbourhood includes closely related pseudogene = 95-98% similarity at sequence level = misalignment during recombination
Patients present with severe hepatosplenomegaly and various skeletal disorders
Can get GBA0 and GBA+ forms of the disease = either complete absence of activity or reduced activity
Misalignment in crossover event thought to cause Gaucher’s Disease
Mutations appear to come from the pseudogene
Enzyme is initially inactive = associates with charged headgroups and becomes partially active
Requires Saposin C as an activator
Saposin C = 80 AA protein - solubilises the substrate = making it more available to the enzyme
Mutation in Saposin C are rare but produce similar symptoms to Gaucher’s Disease
What are the common features of disorders of lysosome related organelles?
Lysosomes biogenesis is an ordered process requiring specific, specialised assembly components
Many specialised cells contain both lysosomes and lysosome-related organelles (LROs):
Melanosomes = pigmentation
Delta granules = found in platelets = blood clotting
Lamellar bodies = found in lungs = produce surfactant
Lytic granules = cytotoxic T lymphocytes
Azurophilic granules
Basophil granules
Defective LRO biogenesis leads to a variety of rare genetic disorders
What are the common features of Hermansky-Pudlak syndrome?
Group of recessive disorders characterised by defects in cell-type specific lysosomal organelles
Defects of components required for formation of organelles
At least eight disease variants in humans (HPS1-8) = different gene products
HPS 1, 3 and 4 show mutations in novel proteins of unknown function
HPS2 = β3A subunit of AP3 adaptin = recognises cargo which becomes part of organelle
Presentations include albinism, prolonged bleeding, pulmonary fibrosis and/or granulomatous colitis (“cobblestone colon”)
In the disease state, there is defective distribution of melanogenic proteins in LRO proteins = clustering of melanophores - don’t distribute properly
In LRO disorders such as CHS (Chediak-Higashi Syndrome), the lysosome-related organelles are larger than is normal
What are the common features of mucopolysaccharidoses?
Inherited disorders leading to accumulation of SULFATED products
Examples include:
Hurler disease
Hunter disease
Scheie disease
Sanfilippo disease
Neufeld first identified that MPS diseases were the result of different discrete individual defects = individual defects - specific mutation
Co-culture of cells with MPS I and MPS II results in corrections of BOTH cell populations to wild-type
What did Neufeld’s co-culture experiments demonstrate?
In normal fibroblasts, you would get synthesis of MPS , secretion of radiolabelled MPS and degradation.
However, in LRO storage diseases there is a buildup of radioactive insoluble material in cells. Co-culturing these cells brings levels of detectable radioactive insoluble material down to a near normal level.
How does enzyme replacement therapy work?
Approach worth pursuing due to HETEROPHAGY = ability to endocytose from the environment = contents part of secondary lysosomes
Normal cells secrete M6P labelled lysosomal enzymes = hydrolytic enzymes
These can be taken up by endocytosis mediated by PM-associated M6P receptor = cation-independent mannose-6-phosphate receptor on cell surface
Lysosomes can pick up enzymes from environment
Neufeld’s and other similar experiments a clue
Pilot studies undertaken in ‘70s on Fabry, Pompe, Sandhoff and Gaucher’s disease
Problems associated with diseases manifesting with severe neurological deficits = not amenable to treatment = can’t cross the blood-brain barrier
Approach: purified human enzymes carrying M6P
Success: Type 1 Gaucher successfully treated by infusion of 2-3mg/kg on alternate weeks
Very little enzyme gets into the cells = doesn’t matter - only a small amount is required to be clinically effective
Over 3000 patients worldwide have been treated this way since 1990
Also use transplantation of donor cells for secretion of missing enzyme = bone marrow can secrete functional cells
How does enzyme activity enhancement therapy work?
ERT successes show that only need “low heterozygote” level to correct pathology
Many LSDs due to protein misfolding = many mutations are point mutations = protein misfolding defects
Hypothesis: a soluble chaperone may prevent misfolding allowing retention of enough activity to prevent disease = promote folding = allows enzyme to get passed ER “quality control”
Proof of principle: Fabry disease (cardiac variety)
X-linked (male expressed)
Results in renal failure, vascular disease of heart and brain, death in 40s/50s
Low activity of α-galactosidase A (1-10% of normal)
Accumulation of glycolipid (globotriaosylceramide; GL3) in secondary lysosomes
In vitro tests with Fabry cardiomyocytes showed increased lysosomal α-galactosidase A
Treatment: galactose infusion (1g/kg, 3 time a week) showed remission of pathology over 3 years = galactose acts a chaperone stabilising the α-galactosidase A
What are some of the other treatments for lysosomal disorders?
Treatment for LRO biogenesis disorders are limited
LRO biogenesis disorder quite often fatal
In Chediak-Higashi Syndrome, this is often due to “accelerated phase” lymphoproliferative problems
In Hermansky-Pudlak Syndrome, it is often due to pulmonary fibrosis
Pirfenidone used to treat pulmonary fibrosis = prevents collagen synthesis = reduces pulmonary scarring
Antibiotics, anti-inflammatory agents (Remicade. steroids) = treating acute immune components of disease
Platelet and haemotopoietic transfusions used successfully in CHS = promote clotting
Give a summary of lysosomal disorders.
Lysosomes are single-membrane bound hydrolytic organelles containing many acid hydrolases
Targeting of lysosomal enzymes mainly via presence of M6P moiety
LSDs are a heterogenous family of disorders involving accumulation of various glycolipid, polysaccharide and peptidoglycan metabolites
LSDs result from mutation of a single primary enzyme
Diseases = progressive; defect = mechanical
Examples include Tay-Sachs, Gaucher, Hunter, Hurler, Fabry, Pompe, Niemann-Pick and MPS I-VII
Prevention by screening/counselling
Cures? - not as such - ERT and EET offer hope for sufferers of some diseases
