The Mitochondrial Genome

Question 1

What are the features of the mitochondrial genome?

Answer 1

Double stranded circular molecule 
Heavy and light strand
Multi copy gene
37 genes
No introns
D loops is a non-coding region where replication and transcription are initiated
Maternally inherited so no recombination

Question 2

What numbers and types of genes make up the 37 mitochondrial genes?

Answer 2

13 OXPHOS protein subunits
22 transfer RNAs
2 ribosomal RNAs

Question 3

What do the OXPHOS proteins consist of?

Answer 3

5 enzyme complexes (CI-CV)

Question 4

What is special about the OXPHOS complex II?

Answer 4

It’s the only entirely nuclear coded subunit

Question 5

What does the non-coding mtDNA region contain?

Answer 5

Regulatory sequences for replication and transcription

Question 6

Where does mtDNA replication start?

Answer 6

The origin of the heavy strand

Question 7

Where does mtDNA transcription start?

Answer 7

Heavy strand and light strand promoters

Question 8

What is mtDNA packaged into?

Answer 8

Nucleoids

Question 9

How many copies of mtDNA do you get per nucleoid?

Answer 9

One or two copies

Question 10

What does transcription factor A act as in mtDNA?

Answer 10

A histone protein

Question 11

What are the exceptions to the universal genetic code?

Answer 11

AUA codes for methionine in mtDNA and isoleucine in nuclear DNA
UGA codes for tryptophan in mtDNA and is a stop codon in nuclear DNA
AGA and AGG are stop codons in mtDNA not argenine

Question 12

What are mtDNA variants called?

Answer 12

Haplogroups

Question 13

Why is the human mtDNA genotype specifically split into haplogroups?

Answer 13

MtDNA is maternally inherited so no recombination occurs

Question 14

Does the mitochondria need both mtDNA and nuclear DNA to function?

Answer 14

Yes

Question 15

How many OXPHOS proteins does the mtDNA encode?

Answer 15

13

Question 16

What is the mtDNA polymerase called?

Answer 16

Polymerase gamma

Question 17

What are the subunits of Polymerase gamma?

Answer 17

One catalytic subunit (A) and two accessory subunits (B)

Question 18

What does the Polymerase gamma catalytic subunit contain and what does it do?

Answer 18

Contains 3’-5’ exonucelase domains to proofread newly synthesised DNA

Question 19

What do the Polymerase gamma accessory subunits contain and what do they do?

Answer 19

Enhance interactions with the DNA template and increases activity and processing of the polymerase gamma A

Question 20

What is the mtDNA helicase called?

Answer 20

TWINKLE

Question 21

How many subunits is TWINKLE made up of?

Answer 21

Six

Question 22

What does the mitochondrial single stranded binding protein do?

Answer 22

Binds to single stranded DNA

Question 23

Why does mitochondrial single stranded binding protein bind to single stranded DNA?

Answer 23

Protects against nucleuses, prevents secondary structure formation and enhances mtDNA synthesis by stimulating TWINKLE helicase activity

Question 24

What is the strand displacement model of mtDNA replication?

Answer 24

Parental heavy strand displaced and coated with MtSSBP
TWINKLE helicase unwinds MtDNA
Mitochondrial RNA polymerase synthesise RNA primer using the light strand as a template
Polymerase gamma uses RNA primer to replicate DNA at the OH

Question 25

What are the steps in mtDNA light strand replication?

Answer 25

Stem loop structure formed MtRNA polymerase synthesises RNA primer using the heavy strand as a template Polymerase gamma uses RNA primer to replicate the light strand DNA

Question 26

What does segregation of mtDNA daughter molecules need?

Answer 26

Topoisomerase IIIa

Question 27

What is the most common form of mitochondrial disease?

Answer 27

OXPHOS disorders

Question 28

What does LHON stand for?

Answer 28

Leber's hereditary optic neuroretinopathy

Question 29

What does KSS stand for?

Answer 29

Kearns-Sayre syndrome

Question 30

What does MELAS stand for?

Answer 30

Mitochondrial Encephalopathy, Lactic Acidosis, and Stroke-like episodes

Question 31

What does MERRF stand for?

Answer 31

Myoclonus epilepsy red ragged fibres

Question 32

What does NARP stand for?

Answer 32

Neurogenic muscle weakness, ataxia, and retinitis pigmentosa

Question 33

What does MINGIE stand for?

Answer 33

Mitochondrial myopathy neuropathy gastrointestinal disease encephalopathy

Question 34

What are the hallmarks of mitochondrial disease?

Answer 34

``` Abnormal bilateral leisions in the basal ganglia of the brain Gamori staining Hypertrophic cardiomyopathy Pseudo-intestinal obstruction Sideroblast anaemia Enlargement of eye vessels ```

Question 35

What is gamori staining?

Answer 35

Clusters of red signal in the muscle fibres

Question 36

How do you diagnose a mitochondrial disease?

Answer 36

Clinical signs Blood and tissue histochemical and analyse measurements Neuro imaging Enzymatic assays of OXPHOS in tissue samples and cultured cells DNA analysis

Question 37

What are some clinical features of mitochondrial disease?

Answer 37

Most have a neurological component and are progressive

Question 38

What types of muscle histology studies are there?

Answer 38

``` Haematoxylon and eosin Gomori trichrome SDH Cox Combined cox/SDH ```

Question 39

What is homoplasmy?

Answer 39

One type of mtDNA in a cell

Question 40

What is heteroplasmy?

Answer 40

One or more mutant mtDNAs in a cell

Question 41

What do heteroplasmy levels determine?

Answer 41

Disease manifestation

Question 42

What % heteroplasmy causes disease manifestation?

Answer 42

80%

Question 43

What do heteroplasmic mutations mean for the inheritance of mutant load?

Answer 43

Random

Question 44

What do homoplasmic mutations mean for the inheritance of mutant load?

Answer 44

Cause the female carriers to transmit to all children

Question 45

How can you identify mtDNA mutations?

Answer 45

Off target reads using WES

Question 46

What do mutations in the mtDNA replication machinery cause?

Answer 46

Secondary mutations in mtDNA

Question 47

What do dominant mutations in TWINKLE cause?

Answer 47

MtDNA deletions and late-onset mitochondrial myopathy

Question 48

What are your options for embryos with mtDNA mutations?

Answer 48

Oocyte donation Prenatal diagnosis Preimplantation genetic diagnosis Mitochondrial replacement therapy

Question 49

What is the most common form of mitochondrial disease?

Answer 49

OXPHOS disorders