Mitochondrial Diseases Flashcards
What are mitochondrial diseases?
Diseases caused by qualitative or quantitative abnormalities of mtDNA
Describe mitochondrial structure
They have an outer membrane and an inner membrane that folds into cristae. Between these = intermembrane space
The matrix is on the inside of the mitochondrion
Mitochondria also contain ribosomes and mitochondrial DNA
Are mitochondria static?
No, they are highly dynamic, undergoing continual fission and fusion
What are the 2 functions all mitochondria can carry out
- The generation of ATP coupled to electron transport (oxidative phosphorylation)
- The expression of an integral genome.
Describe the structure of mtDNA
Closed-circular, double-stranded DNA molecule of ~16.6 kb (heavy strand/light strand)
arranged in nucleoids (nucleoprotein complexes)
Nucleoids are dispersed throughout the mitochondrial network
How does nuclear DNA compare to mtDNA?
Nuclear DNA vs mt: >3 billion base pairs vs 16.6kb 2 copies vs somatic cells have thousands of copies of mtDNA encodes many proteins vs encodes 13 nucleosomes vs nucleoids
How many propteins does mtDNA encode?
13
What do the proteins encoded by mtDNA do?
They are essential subunits of the oxidative phosphorylation enzymes
How are other mitochondrial proteins (not made from mtDNA) made?
They are made by the nucleus and sent into the mt
How many tRNA genes does mtDNA have?
22
What are the proteins involved in oxidative phosphorylation?
Complexes I‒V
Coenzyme Q10
Cytochrome-c
What does each protein complex do in oxidative phosphrylateion
I-II Take electrons from NADH and succinate
III-IV Receive electrons (IV donates the electrons to oxygen, reducing it to wate)
Complex V synthesises ATP using the H+ gradient made by the other complexes
What is another name for complex IV?
Cytochrome C oxidase
No nuclear DNA is needed for the 5 main oxidative phosphorylation complexes. TRUE or FALSE
FALSE
Compare mtDNA replication to nDNA
mtDNA replication occurs at a constant rate throughout the cell cycle and mtDNA continues to be replicated in post-mitotic cells.
nDNA replicates only during S-phase of mitotic cells
How was it found out that mtDNA always replicates independent of cell cycle?
Bromo-deoxyuridine (binds to replicating DNA) was given to cells
Cells which had divided during the experiment had MrdU in their nuclei, however all cells (divided and not) had BrdU staining in their mitochondria
Why are the 2 mtDNA strands called heavy and light?
Two strands of the mtDNA duplex have distinct buoyant densities (“heavy” and “light”) as result of their different base content and can be resolved by equilibrium CsCl gradient centrifugation.
What is the D-loop/displacement loop?
A short, three-stranded structure in which a short nucleic acid strand, complementary to the L-strand, displaces the H-strand
A major control site for mtDNA replication and transcription.
Describe mtDNA replication initiation and the proteins involved
RNA polymerase (POLRMT), TFAM, TFB2M and TEFM make the RNA primer and add it to the heavy origin of replication
What are the functions of TFAM?
transcription factor
packaging role
Describe mtDNA replication
- Twinkle (TWNK) to unwind the DNA duplex
- mitochondrial topoisomerase 1 (TOP1MT) to relax DNA supercoils
- mitochondrial single-stranded binding protein (SSBP1) to stabilize single-stranded regions of mtDNA replicative intermediates
- Synthesis of the mtDNA strands is catalyzed by DNA polymerase gamma
- Once DNA polymerase gamma comes full circle, it encounters the 5’-end of the nascent mtDNA strand. RNA primers are removed by RNase H1 (RNASEH1).
- Further processing of the 5’-end by mitochondrial genome maintenance exonuclease 1 (MGME1).
- gaps are filled in and ligated by DNA ligase III (LIG3)
- the daughter duplexes are topologically linked at the OH region as hemicatenanes, i.e. two circular DNA duplexes bound together via a single-stranded linkage.
- decatenation of the daughter mtDNA molecules is catalyzed by the mitochondrial isoform of topoisomerase 3alpha (TOP3A).
What are the subunits of DNA polymerase gamma?
one POLG subunit and two POLG2 subunits
What does POLG do?
POLG is the catalytic subunit, containing the 5’→3’ DNA polymerase activity as well as a proofreading 3’→5’ exonuclease activity
How is mtDNA inherited?
maternally
Why is mtDNA inheritied maternally?
- The mtDNA copy number in an egg cell (150,000‒700,000) is much higher than in a sperm cell (<10).
- Shortly before or just after fertilisation, sperm mtDNA is largely degraded by endonuclease G.
- After fertilisation, sperm mitochondria are coated with ubiquitin inside the zygote cytoplasm. The ubiquitination selectively earmarks sperm mitochondria for degradation by the proteasomal and autophagosomal systems.
How many proteins are needed for a perfectly functioning mitochondrion?
15,000
What kind of disorders are caused when nuclear genes get involved in depletion of mtDNA in infants?
Alpers disease, myocerebrohepatopathy specturm (MCHS), multi-organ or isolated hepatic disease, myopathy, encephalomyopathy
What causes Alpers disease?
A mutation in the POLG gene leads to mtDNA depletion in the liver, brain and muscle
What causes myocerebrohepatopathy?
A mutation in the POLG, TWNK, or TFAM genes which code for POLG, twinkle and TFAM proteins leads to mtDNA depletion in muscle, brain and liver cells
What kind of inheritance do Alpers disease and Myocerebrohepatopathy have?
autosomal recessive
What are the symptoms of Alpers disease?
Severe and lethal
get a severe encephalopathy, developmental delay, epilepsy, later get liver failure (cause of death)
When is the onset of Alpers disease?
childhood
When is the onset of MCHS?
childhood - die earlier than with alpers
What are the symptoms of MCHS?
Myotonia, myopathy, developmental delay, encephalopathy, liver failure