Lecture 7: Mitochondrial genetic in Health L2 Flashcards
Mitochondria are composed of proteins encoded by two genomes: 2
- Nuclear DNA
- Mitochondrial DNA
Mitochondrial DNA features:
- (mDNA) is found in cell mitochondria
- contains genetic material ONLY FROM THE MOTHER
Nuclear DNA features
- (nuDNA) is found in the CELL NUCLEUS
- Contains genetic material from BOTH PARENTS
Which genome causes mitochondrial diseases..
- Mutations in mtDNA or nuclear DNA genes that make up mitochondria can cause mitochondrial disease
Mitochondrial dysfunction leads to diverse pathologies and diseases….7
1 * Neuromuscular diseases
2 * Dementia and neurological disorders
3 * Diabetes and obesity
4 * Liver disease
5 * Heart disease
6 * Cancer
7 * Ageing
most common genetically inherited metabolic diseases:
Mitochondrial diseases
ORGAN AND MITOCHONDRIAL DNA causes disease = 11
- EYE:
- optic neuropathy
- ophthalmoplegia
- retinopathy - LIVER:
- Hepatopathy - KIDNEY:
- Fanconi’s syndrome
- glomerulpathy - PANCREAS
- Diabetes mellitus - BLOOD
- Pearson’s syndrome - INNER EAR
- Sensorineural hearing loss - COLON
- Pseudo-obstruction - BRAIN
- seizures
- myoclonus
- ataxia
- stoke
- dementia
- migraine - SKELETAL MUSCLE
- weakness
- fatigue
- myopathy
- neuropathy - HEART
- Conduction disorder
- wolf-parkinson-white syndrome
- cardiomyopathy - NUCLEAR DNA —-SUBUNITS —-OXIDATIVE PHOSPHORYLATION —-NUCLEAR DNA
Mitochondrial dysfunction leads to diverse pathologies and diseases - STATISTICS
1 in 8,000 at risk
1 in 15,000 to be affected
Mitochondria are composed of proteins encoded by two genomes
UNDERSTAND SLIDE 5 DIAGRAM AND PROCESS
Mitochondrial DNA mutations and heteroplasmy: 4
- MtDNA mutations have maternal inheritance
2 *Stochastic distribution of mtDNA
3 *Severity of symptoms often can be linked to the
load of mutant mtDNA
4 *Transfer of mutant mtDNA is not well
understood
*Diagnosis of mtDNA mutations in the clinic
relies on: 3
- Sequencing
2 * Biopsies
3 * Enzymatic tests
SLIDE 6 MITOCHONDRIAL INHERITANCE
STUDY FLOW CHART
SLIDE 7 MITOCHONDRIAL…OOCYTE MATURATION AND mtDNA amplification + fertilisation
STUDY FLOW CHART
SLIDE 8 Mitochondrial DNA mutations and heteroplasmy
STUDY FLOW CHART
Mutations in mitochondrial genes cause disease…
Protein-encoding genes = 3
1 * Neurogenic weakness, ataxia and retinitis pigmentosa (NARP) – T8993G/C
2 * Maternally inherited Leigh syndrome (MILS) – T8993G/C
3 * Leber’s hereditary optic neuropathy (LHON) – G3460A, G11778A, T14484C and A14495G
Mutations in mitochondrial genes cause disease
tRNA genes = 3
1 * Mitochondrial encephalopaty with lactic acidosis and stroke-like episodes (MELAS) – A3243G
2 * Myoclonic epilepsy with ragged red fibres (MERRF) – A8344G
3 * Non-syndromic sensorineural deafness – A7445G
Mutations in mitochondrial genes cause disease
rRNA genes = 1
Aminoglycoside-induced non-syndromic deafness – A1555G
Mutations in mitochondrial genes cause disease:
Rearrangements = 3
1 * Chronic progressive external ophthalmoplegia
2 * Kearns-Sayre syndrome
3 * Diabetes and deafness
Mutations in mitochondrial genes cause mitochondrial diseases = 12
- Mutations in mitochondrial genes:
- Frequency 1:4000
3 *Destabilize the secondary or tertiary structure of RNAs
…4 * A3243G MELAS
…5 * A8344G MERRF
6 *Impair the processing of RNAs:
….7 * A3302G cardiomyopathy
…..8 * A5559G
9 *Impair the recognition by tRNA synthetases
…..10 * 5- taurinomethyl-2-thiouridine (τm5 s2 U)
11.*Impair protein synthesis
….12 * 5- taurinomethyl-2-thiouridine (τm5 s2 U)
Mitochondrial Donation
- Eliminating mutant mitochondria = 4
1 *Precludes from transmitting inherited mitochondrial disease to future generations
2 *Already in practice in the United Kingdom
3 *Requires in vitro fertilization
4 *The risks are minimal and research into any
potential risks is carried out
Mitochondrial Donation
Eliminating mutant mitochondria…DIAGRAM
UNDERSTAND FLOW CHART IN SLIDE 11
Gene therapies for
mitochondrial diseases
Modulating the mutant heteroplasmy load: 4
1 *Using protein based therapeutic designed to target only the mutant mtDNA or mtRNA.
2 *Import into mitochondria using existing protein import pathways
3 *CRISPR technologies not possible in mammalian mitochondria as they don’t import RNA
4 *Current gene therapies are under research and development
Gene therapies for
mitochondrial diseases
Modulating the mutant heteroplasmy load…2 FLOW CHARTS ARE IMPORTANT
UNDERSTAND SLIDE 12 DIAGRAMS
Inheritance of mitochondrial diseases caused by mutations in nuclear encoded proteins….
Nuclear mutations follow autosomal dominant or recessive inheritance:
1 *Diagnosis of mitochondrial diseases caused by nuclear DNA mutations in the clinic is
challenging because the diseases are multisystemic and involve multiple different specialists.
2 *There are no treatments or cures to date only
palliative care for patients is available.
Diagnosis of nuclear DNA mutations relies on a
combination of: 3
- Sequencing
- Biopsies
- Enzymatic tests
Inheritance of mitochondrial diseases caused by
mutations in nuclear encoded proteins = SLIDE 12
SLIDE 12
- Autosomal recessive
- Consanguineous autosomal recessive
- autosomal dominant
INTEGRATED APPROACH FOR MITOCHONDRIAL DISEASE DIAGNOSIS …3
- METABOLIC TESTING:
– Blood
– urine
- CSF - Muscle testing
- liver tissue respiromtry
- muscle pathology
- OXPHOSEnzymology
- Protein chemistry
- C0Q10 - GENETIC TESTING:
- mt DNA
- Nuclear DNA
Nuclear mutations in genes encoding mitochondrial proteins…8
- Gene expression defects
* Leigh syndrome – LRPPRC,
TACO1
* Alpers, CMT – POLg
* PEO – Twinkle - Assembly defects
* Leigh syndrome – SURF1
* Cardioencephalomyopathy – SCO2
* Neonatal-onset hepatic failure
and encephalopathy – SCO1
* Leigh and De Toni-FanconiDebre syndrome – COX10 - Homeostasis and import
* Friedreich’s ataxia –frataxin
* Hereditary spastic paraplegia – paraplegin
* Deafness-dystonia syndrome – DDP
* Dominant optic atrophy – OPA1 - Complex I deficiency
* Leigh and Leigh-like syndrome
NDUFS4, NDUFS7 and NDUFS8
* Hypertrophic cardiomyopathy and
encephalomyopathy – NDUFS2
* Macrocephaly, leucodystrophy and
myoclonic epilepsy – NDUFV1 - Complex II deficiency
* Leigh and Leigh-like syndrome - Complex III deficiency
* Encephalopathies, GRACILE
syndrome – BCS1L, UQCRB and UQCRQ - Complex IV deficiency
* Leigh syndrome – COX15, COX8A - Complex V deficiency
* Cardiomyopathy – ATPAF2
Nuclear mutations in genes encoding mitochondrial proteins…DIAGRAM ..understand the process
SLIDE 14
Mitochondrial Diseases affect different organs and have varying severity = 6
- It is important to identify the genetic cause of mitochondrial diseases to
understand how it leads to disease and identify potential treatments.
2 * Some diseases affect many organs and some few
3 * Some diseases are more severe than others
4 * Some diseases occur early in life and some later
5 * A same mutation can result in diverse phenotypes
6 * Different mutations can have a same phenotype
Molecular and functional diagnosis of Mitochondrial Disease = IMPORTANT
KNOW FLOW CHART AND DIAGRAM …SLIDE 16 TO 19
Isolated Complex IV deficiency in a patient with a cytochrome oxidase subunit 1 (CO1) mutation = 5
- Patient - a 10 year old girl
2 *Increased lactic acidosis
3 *Early onset retinitis pigmentosa
4 *Ataxia and peripheral neuropathy
5 *Mutation in CO1 gene coding for a scaffold
protein required for Complex IV activity
Limitations of cell models for mitochondrial disease research = 3
Limitations of cell models for mitochondrial disease research = 3
**Animal models of disease are the best in vivo tool for clinical research
Animal models of disease are the best in vivo tool for clinical research
1 * C. elegans (worms), Drosophila (fruit flies), Mus musculus (mice)
2 * Availability of material, genetic manipulation and physiological relevance
3 * Investigating the disease in the affected tissues
Impaired protein synthesis can generate diverse phenotypes = 7
- Mrps34….Overall impaired translation…ORAGNS ..MITOCHONDRIAL DISEASE
- Taco1…Impaired COXI translation…ORGANS…Mitochondrial Disease
- Ptcd1 Het…Decreased translation… ORGANS …HYPERTROPHIC, CARDIOMYOPATHY, METABOLIC DISEASE
- Ptcd1, Elac2 and Mrpp3 KO…Loss of translation…DILATED CARDIOMYOPATHY
- Mrps12…error-prone translation..ORGANS…Liver Regeneration
- Mrps12….Hyper-accurate translation…ORGANS…DIALTED CARDIOMYOPATHY
- Translation factor…uncoordinated translation…ORGANS….Myopathy and cardiomyopathy
Mitochondrial RNA processing is required for energy production… 4
1.RNase P cleaves the 5’ end of mitochondrial tRNAs and
is composed of 3 subunits: MRPP1, MRPP2 and MRPP3
*All three proteins are required for cleavage
*No requirement for a catalytic RNA
*Mutations in all three proteins cause disease
- *Loss of MRPP3 is embryonic lethal and heart and skeletal muscle KO causes cardiomyopathy and lethality by 11 weeks
- *5′ tRNA processing precedes 3′ tRNA processing
4 *RNA processing links transcription to translation via mitoribosome assembly and this essential for respiration
RNase P cleaves the 5’ end of mitochondrial tRNAs and
is composed of 3 subunits: MRPP1, MRPP2 and MRPP3 = 3
1 *All three proteins are required for cleavage
2 *No requirement for a catalytic RNA
3 *Mutations in all three proteins cause disease
Mitochondrial RNA processing is required
for energy production DIAGRAMS
IMPORTANT ..LOOK AT SLIDE 24
MRPP3 ..KNOW PROCESS AND DIAGRAM
5′ tRNA processing in mitochondria
….RNA sequencing: 3
1 * RNA-Seq enabled capture of tRNAs contained within longer precursor transcripts
2 * PARE enabled capture of 5’ RNA ends
3 * Combined they identified impaired mitochondrial RNA processing
Mutations in mitochondrial RNA processing enzymes cause disease….
…Identified a mutation in MRPP1: 4
- Early onset mitochondrial disease
- Combined OXPHOS defect, multi-systemic disorder, both children died at ~5 months
3 * Decreased stability of the MRPP1 protein
4 * Impaired mitochondrial RNA processing
LRPPRC is a general RNA chaperone required for mRNA stability…
….The leucine-rich pentatricopeptide repeat
cassette (LRPPRC) protein:
1 * A A354V mutation in LRPPRC causes the autosomal recessive French-Canadian variant
Leigh Syndrome
2 * Loss of LRPPRC is embryonic lethal and heart
and skeletal muscle-specific loss of LRPPRC causes dilated cardiomyopathy by 12 weeks
of age in mice
3 * LRPPRC is in a stable complex with SLIRP
that protects it from degradation
4 * The LRPPRC/SLIRP complex is localized in
the mitochondrial matrix where it coordinates
mRNA translation by relaxing their secondary structure
LRPPRC is a general RNA chaperone required for mRNA stability… DIAGRAM
UNDERSTAND AND DRAW
SLIDE 27
Summary: 4
- Mitochondrial diseases are the most common genetically inherited metabolic disorders.
- They can affect young infants as well as adults and the severity of the diseases varies and most often is lethal.
2 * Mitochondrial diseases have several different inheritance modes.
3 * There are no cures for mitochondrial disease so there is an urgent need to find therapeutics and treatments.
- Diagnosis for mitochondrial diseases has improved with
the advances of next generation technologies, however there are still gaps in functional validation of putative pathogenic genes.
4 * There are many different models that can be used to understand the pathology of
mitochondrial diseases and reveal therapeutic targets for drug development
