L27-28: Mitochondrial Diseases

Question 1

Power requirement at rest

Answer 1

100 kcal/hour = 116W

Question 2

O2 consumption at rest

Answer 2

380L/day

Question 3

Brownout

Answer 3

Intentional or unintentional drop in voltage in an electrical power supply system

Question 4

Mitochondrial energy diseases affect ____ births

Answer 4

1/5000

Question 5

True or false: mitochondrial diseases are due to mutation in a single gene.

Answer 5

False, over 200 disease genes.

Question 6

Leigh syndrome

Answer 6

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

Question 7

Endosymbiont Hypothesis

Answer 7

Primitive aerobic bacterium phagocytosed by a fermentative eukaryotic cell. Symbiosis and gradual loss of mtDNA functions.

Question 8

Evidence for Endosymbiont Hypothesis

Answer 8

mtDNA circular, lacks histones.
Inner membrane bacteria-like
rRNA is chloramphenicol sensitive, and emetine resistant

Question 9

How many genes encode mitochondrial proteins?

Answer 9

37 mtDNA genes and 1200-1500 genes encoded by nucleus that encode mitochondrial proteins

Question 10

How many protein coding genes in mtDNA?

Answer 10

13

Question 11

Adult vs. Childhood onset mitochondrial diseases

Answer 11

Adult: 70% maternally inherited mtDNA mutations, autosomal dominant, sporadic
Childhood: 30% maternally inherited mtDNA mutations, mostly autosomal recessive

Question 12

True or false: There are no introns in mitochondrial DNA

Answer 12

True

Question 13

Which OXPHOS complex does not contain subunits of mtDNA?

Answer 13

Complex II

Question 14

Why is mtDNA inherited maternally?

Answer 14

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.

Question 15

Heteroplasmy

Answer 15

Co-existence of mutant and wildtype mtDNA

Question 16

Threshold effect

Answer 16

Minimum proportion of mutant mtDNA needed for each tissue to become dysfunctional

Question 17

mtDNA bottleneck

Answer 17

In early oogenesis, a small number of genomes are selected to repopulate the oocyte, allowing rapid shifts in heretoplasmy

Question 18

True or False: OXPHOS subunits are encoded by both mtDNA and nuclear DNA

Answer 18

True

Question 19

True or false: Each cell has a single copy of mtDNA

Answer 19

False, can have multiple copies of mtDNA per cell

Question 20

mtDNA has a ____ mutation rate, compared to nuclear genes

Answer 20

high

Question 21

How common are OXPHOS disorders?

Answer 21

1/5000 births

The most common neurogenetic disease

Question 22

Challenges of OXPHOS molecular diagnosis

Answer 22

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

Question 23

Sanger vs. NextGen Sequencing

Answer 23

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

Question 24

Human genome contains how many genes?

Answer 24

~20,000

Question 25

Human genome contains how many bp?

Answer 25

3Gb (3x10^9 bp)

Question 26

Illumina sequencing by synthesis

Answer 26

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)

Question 27

Ion torrent sequencing

Answer 27

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.

Question 28

Oxford Nanopore MinION sequencing

Answer 28

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.

Question 29

Rare diseases affect 1 in ___ people

Answer 29

2000