Mitochondrial Genetics Course Notes PDF

Mitochondrial Genetics: inheritance, disease and complex phenotypes Alan Hodgkinson  Learning Objectives 1. To understand the function of mitochondrial DNA (mtDNA) and the processes that drive variation in mtDNA across cells, individuals and tissues. 2. To identify t...

Mitochondrial Genetics: inheritance, disease and complex phenotypes Alan Hodgkinson  Learning Objectives 1. To understand the function of mitochondrial DNA (mtDNA) and the processes that drive variation in mtDNA across cells, individuals and tissues. 2. To identify the role of the nuclear genome in modulating mitochondrial processes. 3. To infer the impact of nuclear-mitochondrial relationships on mitochondrial diseases. 4. To consider the role of mitochondria in common diseases.  Mitochondrial Genetics 1. The genetics of mitochondrial inheritance and variation 2. Mitochondrial disorders: the interplay between nuclear and mitochondrial genomes 3. The influence of mitochondrial variation on complex disease Nuclear Genome Nuclear DNA: 23 pairs of chromsomes:  Nuclear Genome Father Mother Each chromosome has two copies: one Offspring inherited from the mother and one from the father.  Nuclear Genome Disease Mutation Father Mother Disease causing mutations can be Offspring inherited through generations under various different dominance models. Eukaryotic Cell Mitochondria Apoptotic Cell Death Intracellular Signalling Lipid Metabolism Thermogenesis + Cellular Energy Production! OXPHOS – ATP Production Taken from: Werner et al (2012) NEJM 306:1132-41 OXPHOS – ATP Production  Mitochondrial DNA Mitochondria have their own DNA!  Mitochondrial DNA DNA is circular and contains genes coding for protein of the OXPHOS chain Taken from: Taylor et al (2005) Nature Reviews Genetics 6: 389-402 Mitochondrial DNA 13 Polypeptides 22 transfer RNAs 2 ribosomal RNAs Taken from: Zeviani et al (2004) Brain 127: 2153-2172  Mitochondrial DNA DNA is circular and contains genes coding for protein of the OXPHOS chain Taken from: Taylor et al (2005) Nature Reviews Genetics 6: 389-402  Mitochondrial DNA Each mitochondria contains multiple copies of DNA Taken from: Taylor et al (2005) Nature Reviews Genetics 6: 389-402  Mitochondrial DNA Common mitochondrial genome Each mitochondria contains multiple copies of DNA  Mitochondrial DNA Common mitochondrial genome Homoplasmy Each mitochondria contains All copies are identical multiple copies of DNA (can also refer to a position in the genome)  Mitochondrial DNA Common mitochondrial genome Homoplasmy Heteroplasmy Each mitochondria contains All copies are identical More than one type of multiple copies of DNA (can also refer to a MT genome present (or position in the genome) variation at position)  Mitochondrial DNA Father Mother Mother Offspring Offspring Mitochondrial vs Nuclear Genomes Taken from: Taylor et al (2005) Nat Rev Gen 6:389-402  Nuclear and Mitochondrial genomes Proteins encoded in the nuclear genome are important for mitochondrial function: Nuclear derived proteins form part of the OXPHOS chain: Taken from: Schon et al (2012) Nat Rev Gen 13:878-890  Nuclear and Mitochondrial genomes Nuclear derived proteins involved in the replication and repair of the mitochondrial genome:  Nuclear and Mitochondrial genomes Nuclear derived proteins control transcription and processing of mitochondrial genome...as well as being involved in translation. From: Rackham et al (2012). WIREs RNA. 3, 675–695  Nuclear and Mitochondrial genomes Nuclear derived proteins control transcription and processing of mitochondrial genome...as well as being involved in translation. Nuclear gene products! From: Rackham et al (2012). WIREs RNA. 3, 675–695  Nuclear and Mitochondrial genomes Nuclear derived proteins control transcription and processing of mitochondrial genome...as well as being involved in translation. Nuclear and Mitochondrial genomes interact and both are essential for cellular energy production From: Rackham et al (2012). WIREs RNA. 3, 675–695  The Big Picture Each cell can contain as many as 10000 Mitochondria Cellular Energy Production Each mitochondria may contain 10 circular genomes – creating variation! Nuclear gene products are Nuclear genes play a role in part of OXPHOS chain and are replication, transcription and involved in many processes in translation of mitochondrial the mitochondria. genome Nuclear Genome  The Big Picture Each cell can contain as many as 10000 Mitochondria Cellular Energy Production Mitochondrial diseases and disorders can have both nuclear and mitochondrial origins Each mitochondria may contain 10 circular genomes Nuclear gene products are Nuclear genes play a role in part of OXPHOS chain and are replication, transcription and involved in many processes in translation of mitochondrial the mitochondria. genome Nuclear Genome  Mitochondrial Disorders General Features: Respiratory chain deficiency Reduced cellular oxygen consumption and ATP synthesis. Increased lactic acid in blood and cerebral fluid. Overproduction of reactive oxygen species (ROS).  Mitochondrial Disorders Mitochondrial disorders may manifest in multiple systems and tissues  Mitochondrial Disorders General Features: Respiratory chain deficiency Reduced cellular oxygen consumption and ATP synthesis. Increased lactic acid in blood and cerebral fluid. Overproduction of reactive oxygen species (ROS). May manifest in multiple systems and tissues. More likely to influence tissues with higher energy demand (brain/heart/kidney/skeletal muscle). 1. Nuclear origin 2. Nuclear origin with mitochondrial dysfunction 3. Mitochondrial origin  Mitochondrial Disorders 1. Mitochondrial disease caused by nuclear genome mutations: Follow same inheritance patterns as Mendelian disorders. Can be caused by defects to genes involved in replication, transcription, translation and repair of MT DNA.  Mitochondrial Disorders 1. Mitochondrial disease caused by nuclear genome mutations: Follow same inheritance patterns as Mendelian disorders. Can be caused by defects to genes involved in replication, transcription, translation and repair of MT DNA. Example: Friedreich Ataxia - Inherited neurodegenerative disorder, Incidence ~1/30,000 - Progressive gait and limb ataxia - Unstable GAA expansion in FXN gene – iron trafficking in mitochondria - Autosomal recessive.  Mitochondrial Disorders 2. Mitochondrial disease caused by nuclear genome mutations, with mitochondria dysfunction dependence: Defects in genes that monitor and regulate mitochondria. Requires mitochondrial dysfunction. Important in ageing.  Mitochondrial Disorders 2. Mitochondrial disease caused by nuclear genome mutations, with mitochondria dysfunction dependence: Defects in genes that monitor and regulate mitochondria. Requires mitochondrial dysfunction. Important in ageing. Example: Parkinson’s Disease - Affects 1-2% of the population. - Rigidity, postural instability and tremor: stem from progressive loss of dopaminergic neurons in brain. - Two genes identified: Parkin and PINK1.  Mitochondrial Disorders Example: Parkinson’s Disease Two genes identified: Parkin and PINK1. Normally, PINK1 imported into mitochondria, cleaved and released Damage to MT caused depolarisation of membranes. PINK1 accumulates in outer membrane of MT. Parkin recruited to mitochondria and triggers autophagy. Damage to this pathway leads to build-up of dysfunctional mitochondria From: Pickrell et al (2015). Neuron review. 85, 257–273  Mitochondrial Disorders 3. Mitochondrial disorders caused by mutations in mitochondrial genome Caused by point mutations or re-arrangements. Similar general features to disorders caused by nuclear mutations. Inherited from mother. >500 sites linked to disease, high percentage (over 50%) in tRNAs.  Mitochondrial Disorders 3. Mitochondrial disorders caused by mutations in mitochondrial genome Caused by point mutations or re-arrangements. Similar general features to disorders caused by nuclear mutations. Inherited from mother. >500 sites linked to disease, high percentage (over 50%) in tRNAs. Example: Leber’s hereditary optic neuropathy (LHON) Most common MT disorder. Sub-acute loss of central vision in young adults. Predominantly affects men. Caused by homoplasmic mutations in one of three genes: - 11778G->A in NADH dehydrogenase 4 (ND4) - 3460G->A in ND1 - 14484T->C in ND6  Threshold Model Mutations causing disease may be heteroplasmic and have a threshold effect: Low level heteroplasmy No effect  Threshold Model Mutations causing disease may be heteroplasmic and have a threshold effect: Low level heteroplasmy Medium level heteroplasmy No effect Milder symptoms Example: 8993T->G in ATP6 At 70-90%: Neuropathy, ataxia and retinitis pigmentosa (NARP). Late acting, debilitating, non- fatal  Threshold Model Mutations causing disease may be heteroplasmic and have a threshold effect: Low level heteroplasmy Medium level heteroplasmy High level heteroplasmy No effect Milder symptoms More severe symptoms Example: 8993T->G in ATP6 Example: 8993T->G in ATP6 At 70-90%: Neuropathy, ataxia At >90%: Maternally inherited and retinitis pigmentosa (NARP). Leigh’s Syndrome (MILS). Late acting, debilitating, non- Fatal encephalopathy fatal  Continuous Clinical Spectrum Mother (I-2), has 31% of mutant mtDNA in her blood is asymptomatic  Continuous Clinical Spectrum Mother (I-2), has 31% of mutant mtDNA in her blood is asymptomatic Youngest brother (II-3), carrying 94% mutant load, is severely affected with Leigh Disease  Continuous Clinical Spectrum Mother (I-2), has 31% of mutant mtDNA in her blood is asymptomatic Youngest brother (II-3), carrying 94% mutant load, is severely affected with Leigh Disease Older brother (II-2), with 80%, has NARP syndrome  Continuous Clinical Spectrum Mother (I-2), has 31% of mutant mtDNA in her blood is asymptomatic Youngest brother (II-3), carrying 94% mutant load, is severely affected with Leigh Disease Older brother (II-2), with 80%, has NARP syndrome What is going on here?  Mutations in MT Genome Conundrum: Mitochondria related diseases: 1 in 5,000 individuals affected But a much higher frequency of individuals carry a mutation associated with MT disease Compared 3168 matched neonatal-cord-blood and maternal blood samples 1 in 200 people carry at least one of the 10 most common pathogenic MT mutations. Elliott et al (2008). AJHG. 83, 254–260  Mutations in MT Genome Conundrum: Mitochondria related diseases: 1 in 5,000 individuals affected But a much higher frequency of individuals carry a mutation associated with MT disease Looked at whole MT sequences from >1000 individuals (1000G Project). ~90% of people carry at least one heteroplasmy At least 20% of people carry a mutation associated with disease. Heteroplasmies with >60% presence have lower predicted effects on function Ye et al (2014). PNAS. 29, 10654–659  Mutations in MT Genome Conundrum: Mitochondria related diseases: 1 in 5,000 individuals affected But a much higher frequency of individuals carry a mutation associated with MT disease Complexity of inheritance, origin, detection and prevalence make MT diseases challenging to study and diagnose Looked at whole MT sequences from >1000 individuals (1000G Project). ~90% of people carry at least one heteroplasmy At least 20% of people carry a mutation associated with disease. Heteroplasmies with >60% presence have lower predicted effects on function Ye et al (2014). PNAS. 29, 10654–659  Strategy for Mitochondrial Diseases Taken From: Tanaka et al Nature Medicine 19, 1578–1579 (2013) Oocytes from a patient carrying mitochondria with mtDNA mutations (gray) and a healthy egg donor with normal mitochondria (purple) are retrieved. The resulting oocyte contains normal can undergo IVF to produce an embryo.  Strategy for Mitochondrial Diseases Nuclear genome editing tools cannot be targeted to mitochondria New: cytidine deaminase enzyme called DddA, catalyses conversion of cytosine to uracil. Conversion without inducing double-strand DNA breaks: suited to editing the mitochondrial genome, which lacks efficient mechanisms for repair of these Taken From: Aushev et al Nature 583, 521-522 (2020)  Mitochondrial and Common Disease Some MT mutations cause what are considered to be complex diseases, but only in rare cases and affecting small numbers of people (e.g. diabetes). Associations between mitochondrial variants and complex disease are gaining support – mostly homoplasmies and large scale changes Susceptibility to complex disease: examples of Alzheimer’s disease, Huntington’s disease, metabolic syndrome, myocardial infarction, Parkinson’s disease and type 2 diabetes. Most links are correlative, not validated and difficult to prove functionally.  Mitochondrial and Common Disease Cancer Warburg found that tumours produce excess lactate in the presence of oxygen – aerobic glycolysis: ‘Warburg Effect’ High rate of glucose uptake by solid tumours is used for diagnosis – FDG accumulation detected by positron emission tomography. Since then, much work focussed on finding mechanisms for defective mitochondria in cancer cells.  Mitochondria and Cancer Theory supported by idea that impairment of OXPHOS function stimulates AKT cell survival pathway = down regulation of apoptosis. Increased number of mitochondrial mutations in tumour cells. Mt-ND3 mutation associated with breast cancer (Canter et al, 2005) Mt-CO1 mutation associated with prostate cancer (Petros et al 2005) Are these mutations linked to mitochondrial function or involved in tumorigenesis?  Mitochondria and Cancer Mouse model (and patients) with mt-ND5 mutation: Tumor growth not altered but Metabolic and immune state of tumors altered: From: Mahmood et al (2023). bioRxiv 2023.03.21.533091  Mitochondria and Cancer Removal of mtDNA from cancer cells -> reduced tumour formation (Weinburg et al 2010). Cells without mtDNA show slower tumour formation Metastatic cells acquire mtDNA from host cells to re- establish tumour-initiating efficiency. From: Tan et al (2015). Cell Metabolism. 21, 81-94  Mitochondria and Cancer Removal of mtDNA from cancer cells -> reduced tumour formation (Weinburg et al 2010). Cells without mtDNA show slower tumour formation Metastatic cells acquire mtDNA from host cells to re- establish tumour-initiating efficiency. Functional Mitochondria are important! From: Tan et al (2015). Cell Metabolism. 21, 81-94  Mitochondria and Cancer Removal of mtDNA from cancer cells -> reduced tumour formation (Weinburg et al 2010). Cells without mtDNA show Tumours undergo a change in bio- slower tumour formation energetic profile associated with Metastatic cells acquire mtDNA from host cells to re- mitochondrial processes: establish many tumour-initiating causal mechanisms leftefficiency. to explore! Functional Mitochondria are important! From: Tan et al (2015). Cell Metabolism. 21, 81-94  Summary Mitochondria have their own DNA that contributes gene products of OXPHOS system, generating cellular energy. Nuclear genes are involved in this system and also play a role in the regulation and processing of the mitochondrial genome. As a result, mitochondrial disorders are often complex and difficult to understand. As mitochondria are present in almost every cell, mitochondrial diseases are wide ranging and affect multiple systems: Mendelian and complex. The threshold effect governs the severity of disorder through varying levels of heteroplasmy. Mitochondrial replacement therapy may reduce incidence of disease, but ethical and biological implications need to be understood. The role of mitochondria in complex disease is highly controversial. Clear effects in Parkinson’s disease, implicated in cancer among others. Underlying mechanisms of disease mostly unknown.