Mitochondrial Diseases Flashcards
mitochondrial proteome
encompasses ~1130 proteins
- majority encoded by nuclear DNA and imported into mitochondria following their cytosolic translation
- 13 polypeptides encoded by mitochondrial DNA
mitochondrial genome
- 5-10 copies of mtDNA per mitochondrion
- thousands of copied of mtDNA per cell
- required for translfating the transcripts of mitochondria encoded polypeptides
- completely maternally inherited
- > 200 different rearrangements and missense variants in mtDNA causing human disease
size of mitochondrial genome
16 569 bp
structure of mitochondrial genome
double stranded closed circle
encodes 37 genes
- 13 subunits of enzymes involved in ox phosphorylation
- two rRNA and 22 tRNA genes
> necessary RNA components for intra-mitochondrial protein synthesis
OH and OL are the original heavy- and light-strand mtDNA replication
replicative segregation
each cell mitochondria replicate and sort randomly
mitochondria are distributed randomly between two daughter cells
significant variability in manifestations of mitochondrial disorders among different tissues and/or patients
homoplasmy and heteroplasmy
random distribution of mutant and normal mtDNA into daughter cells and organelles
different proportion of wild-type and mutant mitochondrial genomes
mixture of mutant and wild-type mtDNA in organelles called heteroplasmy
if pure normal or pure mutated mtDNA = homoplasmy
mitochondrial genome bottleneck
- number of mtDNA molecules within developing oocytes is reduced before being amplified to the huge total seen in mature oocytes
- the restriction and subsequent amplification of mtDNA during oogenesis = bottleneck
- mothers with high proportion of mutant mtDNA = more likely to produce eggs with higher proportion of mutant mtDNA = more likely affected offspring
T or F. Primary mitochondrial diseases can affect any organ and any system
T! but mostly bain and muscles
or usually multi-organ affected
which organs are affected the most with primary mitochondrial diseases
high energy organs = brain and muscles
pattern of inheritance if multiple family members affected
mitochondrial inheritance
autosomal recessive inheritance
autosomal dominant
X-linked inheritance
this is helpful in muscle biopsies but cannot differentiate between primary and secondary mt disorders
histochemical stains
functions of the mitochondria
ATP synthesis via ox phosphorylation system
contribution to regulation of phospholipid biosynthesis,
reactive O2 species (ROS) signalling, Ca2+ handling, iron-sulfur cluster homeostasis
serv as signalling platforms for several pathways:
- innate immune response
- programmed cell death through apoptosis
- cell differentiation
mitochondrial DNA synthesis
- continuous event
- not regulated by cell cycle
- require balanced supply of intra-mitochondrial nucleotides imported from cytosol via transporters
- balanced quantities of enzymes through mitochondrial fission and fusion
- all proteins required for synthesis encoded by nuclear genes
- pathogenic variants in those nuclar genes impair mtDNA synthesis
> quant defects = mtDNA depletion
> qual defects = mtDNA deletions
mitochondrial DNA maintenance
- essential to functioning of mitochondria
- meeting energy needs of all cells
- requires proteins essential for mtDNA synthesis, maintenance of mitochondrial nucleotide pool, mediating mitochondrial fusion
cell and tissue dysfunction results when the fraction of mitochondria carrying a mutation exceeds a _______ level
threshold
replicative segregation of a heteroplasmic mitochondrial mutation
T or F. Females heteroplasmic for missense variants or duplications will pass those to all of their children
T! BUT severity can vary in their children
- depends on fraction of mutant variant in their mother
- due to random chance operating small numbers of mitochondria per cell at oocyte bottleneck
T or F. Heteroplasmic deletions, duplications and missense variants are heritable
F! deletions are not heritable
what are primary mitochondrial diseases due to?
nuclear or mitochondrial genome variants
inheritance patterns of primary mitochindrial diseases
mitochondrial
aut recessive (most common)
autosomal dom
maternal
X-linked
sporadic
nuclear DNA genes
more than 1000 (compared to 37 mtDNA genes)
if only one family member is affected or simplex case
aut dom (de novo or decreased penetrance)
aut recessive, X-linked, maternally inherited
acquired (non-genetic)
childhood onset of encephalomyopathy and adults with PEO or KSS represent these
simplex cases
multiple affected family members
mitochondrial inheritance
aut recessive inheritance
aut dom inheritance
X-linked inheritance
molecular genetic investigations for mitochondrial diseases
- mit disorders multigene panels for specific phenotypes/neuroimaging feats
- exome sequencing
- genome sequencing
- mit genome sequencing
> mtDNA pathogenic variants are heteroplasmic, proportion of mutated mtDNA may be undetectable in blood
> use skeletal muscle, urinary epithelium, etc.
if no specific features in mit disorders
exome sequencing
some common mtDNA pathogenic variants (eg. large-scale deletions cause CPEO) may only be detected here
skeletal muscles
supportive investigations for mitochondrial diseases
plasma/CSF lactate/ pyruvate measurements
cardiac assessments
neuroimaging (brain CT/MRI)
magnetic resonance spectroscopy
tissue biopsy (muscle, skin, liver) to investigate ragged fibers, ox phosphorylation enzyme activity measurements, muscle histopathology/chemistry, muscle electron microscopy
neurophysiological studies (EEG, EMG, NCV)
how can we histologically asses mitochondrial function?
H&E
modified Gomori trichrome = ragged red fibres (RRF) due to abnormal subsarcolemmal proliferation of mitochondria
muscle fiber diameter (hypertrophic fibers)
presence of any abnormal inclusions or central nuclei
lacking histocytochemical cytochrome c oxidase (COX) activity = COX deficient fibers
T or F. Histochemistry for muscle biopsy needs to be requested
F! It’s routine
T or F. Histochemistry can distinguish between 1ry and 2ry mt disorders
F!
why is prenatal molecular genetic testing and interpretation for mtDNA disorders difficult?
mtDNA heteroplasmy
- percentage level of mutated mtDNA in a chorionic villus sampling biopsy may not reflect percentage level of mutated mtDNA in other fetal tissues
- percentage level of mutated mtDNA may change during development and throughout life
successful prenatal molecular diagnosis has been carried out for these pathogenic variants using DNA extracted from fetal cells via amniocentesis /CVS
m.8993T>G
m.8993T>C
- more ven tissue distribution
- percentage level of these 2 variants does not appear to change significantly overtime
- 100% homoplasmy
SMD vs PMD
SMD can be inherited or acquired
PMD (primary) = only inherited
chronic progressive ophthalmoplegia
- ptosis, external opthalmoplegia
- single large-scale mtDNA deletion (1.1-10kb)
- m.3243A>G
- at least 35 distinct mtDNA missense variants
- at least 12 nuclear DNA pathogenic variants
- one phenotype and multiple genotypes
