L27-28: Mitochondrial Diseases Flashcards
Power requirement at rest
100 kcal/hour = 116W
O2 consumption at rest
380L/day
Brownout
Intentional or unintentional drop in voltage in an electrical power supply system
Mitochondrial energy diseases affect ____ births
1/5000
True or false: mitochondrial diseases are due to mutation in a single gene.
False, over 200 disease genes.
Leigh syndrome
Disorder of mitochondrial oxidative phosphorylation
Typically healthy until 6 months
Progressive episodic neurodegeneration
High plasma lactic acid, death usually by 3 years of age
Diagnosis reqires an invasive muscle biopsy
No effective therapy and genetic diagnosis often unattainable
Endosymbiont Hypothesis
Primitive aerobic bacterium phagocytosed by a fermentative eukaryotic cell. Symbiosis and gradual loss of mtDNA functions.
Evidence for Endosymbiont Hypothesis
mtDNA circular, lacks histones.
Inner membrane bacteria-like
rRNA is chloramphenicol sensitive, and emetine resistant
How many genes encode mitochondrial proteins?
37 mtDNA genes and 1200-1500 genes encoded by nucleus that encode mitochondrial proteins
How many protein coding genes in mtDNA?
13
Adult vs. Childhood onset mitochondrial diseases
Adult: 70% maternally inherited mtDNA mutations, autosomal dominant, sporadic
Childhood: 30% maternally inherited mtDNA mutations, mostly autosomal recessive
True or false: There are no introns in mitochondrial DNA
True
Which OXPHOS complex does not contain subunits of mtDNA?
Complex II
Why is mtDNA inherited maternally?
A mature human oocyte has 200,000 copies of mtDNA, as opposed to 50 copies in sperm. In a mature human oocyte, half the DNA is mtDNA. As well, the male mitochondria of the sperm are destroyed inside the egg as they are targeted by ubiquitin.
Heteroplasmy
Co-existence of mutant and wildtype mtDNA
Threshold effect
Minimum proportion of mutant mtDNA needed for each tissue to become dysfunctional
mtDNA bottleneck
In early oogenesis, a small number of genomes are selected to repopulate the oocyte, allowing rapid shifts in heretoplasmy
True or False: OXPHOS subunits are encoded by both mtDNA and nuclear DNA
True
True or false: Each cell has a single copy of mtDNA
False, can have multiple copies of mtDNA per cell
mtDNA has a ____ mutation rate, compared to nuclear genes
high
How common are OXPHOS disorders?
1/5000 births
The most common neurogenetic disease
Challenges of OXPHOS molecular diagnosis
Large number of candidate genes
Mostly private mutations - haven’t been found in any other patient
Common mutations are common in only a few genes
Genotype/phenotype correlation often poor
Sanger vs. NextGen Sequencing
Sanger sequencing: 1 target DNA, average of all DNAs,~800 bp per run
NextGen sequencing: Millions of DNAs at a time, single molecule (cluster) DNA sequences, 1.5x10^12bp per run
Human genome contains how many genes?
~20,000
Human genome contains how many bp?
3Gb (3x10^9 bp)
Illumina sequencing by synthesis
1) DNA is fragmented (usually to 150bp) and adapters are added which allow DNA to bind to flowcell
2) Amplified in clusters in a flow cell
3) Sequenced using fluorescent reversible terminator bases (A-T-C-G added to PCR mix)
Ion torrent sequencing
DNA is fragmented and captured onto beads. These beads are then emulsified and amplified - beads are put into PCR mix with oil, and mixed so each bead bound with DNA is within a droplet of oil. The PCR is then performed within the droplet of oil. When the nucleotide incorporates into the DNA, H ion is released.
Oxford Nanopore MinION sequencing
3rd geneneration sequencing technology, MinION is a device single molecule analysis system, designed for single use and to work through the USB port of a computer. Nanopore technology is capable of reads >100 kb. Used in research as read error rate is higher than competitors.
Rare diseases affect 1 in ___ people
2000
