The mitochondrial genome

Question 1

How is ATP generated in the mitochondria?

Answer 1

Generation of ATP by oxidative phosphorylation

Question 2

What are mitochondrial functions?

Answer 2

• Haem synthesis
• Neurotransmitter synthesis
• Nucleotide synthesis

Question 3

What is the structure of the mitochondrial genome?

Answer 3

Double stranded circular molecule

Question 4

What does the mitochondrial genome consist of and not have?

Answer 4

Consists of heavy and light strands and has no introns

Question 5

How many genes does the mitochondrial genome have?

Answer 5

Has 37 genes

Question 6

What are the genes present in the mitochondrial genome and the amount present?

Answer 6

• 13 Oxidative phosphorylation protein subunits
• 22 tRNAs
• 2 rRNAs

Question 7

What is the D loop in the mitochondrial genome?

Answer 7

The D loop is a non-coding region where replication and transcription are initiated

Question 8

How is the mitochondrial genome inherited?

Answer 8

Maternally inherited, no recombination

Question 9

What does the mitochondrial genome encode?

Answer 9

Encodes proteins of oxidative phosphorylation

Question 10

What does the non-coding region contain?

Answer 10

Contains regulatory sequences for replication and transcription

Question 11

Where does mtDNA replication start?

Answer 11

mtDNA replication starts in the origin of heavy strand (OH)

Question 12

Where does transcription of mtDNA start?

Answer 12

Transcription starts at the heavy strand promoter(HSP) and the light strand promoter(LSP)

Question 13

What is mtDNA packaged into?

Answer 13

Packaged into nucleotides

Question 14

How many copies of mtDNA are there per nucleotide?

Answer 14

One or two copies of mtDNA per nucleoid

Question 15

What does transcription factor A(TFAM) act as when mtDNA is packaged?

Answer 15

Acts as a histone protein

Question 16

Genetic code in vertebrate mitochondria DNA vs nuclear DNA

Answer 16

• AUA and AUG code for methionine in mitochondria; whereas in nuclear DNA, AUA codes for isoleucine
• UGA codes for tryptophan in mitochondria; whereas in nuclear DNA, it’s a stop codon
• AGA and AGG are stop codons in mitochondria; whereas in nuclear DNA, it’s arginine

Question 17

What is the endosymbiotic theory of the origin of the mitochondria?

Answer 17

-Mitochondria and chloroplasts originated as bacterial cells that came to live inside larger cells

Question 18

What does the mitochondria require both of?

Answer 18

Requires both nuclear and mtDNA encoded proteins

Question 19

How many proteins does oxidative phosphorylation require?

Answer 19

Oxidative phosphorylation requires >100 proteins but mtDNA only encodes 13 of those proteins

Question 20

What must happen in order to make the 13 proteins for oxidative phosphorylation?

Answer 20

In order to make the 13 proteins for oxidative phosphorylation, protein mtDNA must be:

• Replicated
• Transcribed
• Translated

Question 21

What are all the proteins involved in replication, transcription and translation of mtDNA encoded by?

Answer 21

All proteins involved in replication, transcription and translation of mtDNA are encoded by nuclear genes and imported into mitochondria

Question 22

What are all the other proteins involved in oxidative phosphorylation made by?

Answer 22

All other proteins involved in oxidative phosphorylation are made by nuclear genes

Question 23

What is the structure of the mtDNA polymerase(polymerase gamma)?

Answer 23

• Heterotrimer protein
  
  • One catalytic subunit (POLA)
  • Two accessory subunits (POLB)

Question 24

What does POLgammaA contain?

Answer 24

Contains 3’-5’ exonuclease domain to proofread newly synthesized DNA

Question 25

What does POLgammaB enhance?

Answer 25

Enhances interactions with DNA template and increases activity and processability of POLgammaA

Question 26

What is the structure of mtDNA helicase(TWINKLE)?

Answer 26

TWINKLE: | -Six TWINKLE subunits

Question 27

What does mtDNA helicase, TWINKLE, do?

Answer 27

It unwinds double stranded mtDNA template to allow replication by POLgamma

Question 28

What does the mitochondrial single stranded binding protein bind to and what does it do?

Answer 28

Binds to single stranded DNA - Protects against nucleases - Prevents secondary structure formation - Enhances mtDNA synthesis by stimulating TWINKLE helicase activity

Question 29

Steps involved in the replication of mtDNA

Answer 29

- Replication of heavy strand - Replication of light strand - Replication of both strands complete and segregation of daughter molecules occurs

Question 30

Replication of heavy strand

Answer 30

- Parental heavy strand displaced and coated with mtSSBP - TWINKLE helicase unwinds mtDNA - Mitochondrial polymerase (POLRMT) synthesizes RNA primer using light strand as template - POLgamma uses RNA primer to replicate DNA at OH

Question 31

Replication of light strand

Answer 31

- Heavy strand replication passes OL - Stem loop structure is formed preventing mtSSBP binding - Mitochondrial polymerase (POLRMT) synthesizes RNA primer using heavy strand as a template - POLgamma uses RNA primer to replicate light strand DNA at OL

Question 32

Completion of both strands

Answer 32

- Synthesis proceeds until both strands are fully replicated | - After replication, daughter molecules are segregated

Question 33

Heavy strands

Answer 33

Heavy strands have a high composition of A and G (heavy bases)

Question 34

Light strands

Answer 34

Light strands have a high composition of C and T(light bases)

Question 35

Mitochondrial diseases

Answer 35

- Rare monogenic diseases - Affect highly metabolic organs - Can affect one or several organ systems - Genetically heterogeneous

Question 36

What causes mitochondrial disease?

Answer 36

Mutation in >300 nuclear genes cause mitochondrial disease, not just mtDNA

Question 37

What determines mitochondrial disease manifestation?

Answer 37

Heteroplasmy levels determine mitochondrial disease manifestation

Question 38

How is mtDNA inherited and mutation load?

Answer 38

mtDNA is maternally inherited and inheritance of mutation load is random

Question 39

What way can be used to identify mtDNA mutations

Answer 39

mtDNA mutations can be identified by next generation sequencing