Mitochondria: Impaired mitochondrial translation in human disease

1Organelle factsMitochondrial protein synthesis requires several mitochondrial and nuclear-encoded factors for optimal translation.The clinical presentation of diseases due to defective mitochondrial protein synthesis is very variable and tissue specific presentations are common.The reasons behind the tissue specificity are largely unknown.Besides mitochondrial tRNA mutations and mtDNA deletions or depletion, autosomal recessive mutations have been reported in genes encoding ribosomal proteins, ribosome assembly proteins, mitochondrial aminoacyl-tRNA synthetases, tRNA modifying enzymes and initiation, elongation and termination factors of translation.Frequent and clinically recognisable genetic causes of human diseases due to impaired mitochondrial translation are caused by mutations in mitochondrial tRNA synthetase and tRNA modifying genes.The potential interaction between cytosolic and mitochondrial translation requires further investigations.

2Organelle factsMitochondrial protein synthesis requires several mitochondrial and nuclear-encoded factors for optimal translation.The clinical presentation of diseases due to defective mitochondrial protein synthesis is very variable and tissue specific presentations are common.The reasons behind the tissue specificity are largely unknown.Besides mitochondrial tRNA mutations and mtDNA deletions or depletion, autosomal recessive mutations have been reported in genes encoding ribosomal proteins, ribosome assembly proteins, mitochondrial aminoacyl-tRNA synthetases, tRNA modifying enzymes and initiation, elongation and termination factors of translation.Frequent and clinically recognisable genetic causes of human diseases due to impaired mitochondrial translation are caused by mutations in mitochondrial tRNA synthetase and tRNA modifying genes.The potential interaction between cytosolic and mitochondrial translation requires further investigations.

3Mitochondrial protein synthesis requires several mitochondrial and nuclear-encoded factors for optimal translation.

4The clinical presentation of diseases due to defective mitochondrial protein synthesis is very variable and tissue specific presentations are common.

5The reasons behind the tissue specificity are largely unknown.

6Besides mitochondrial tRNA mutations and mtDNA deletions or depletion, autosomal recessive mutations have been reported in genes encoding ribosomal proteins, ribosome assembly proteins, mitochondrial aminoacyl-tRNA synthetases, tRNA modifying enzymes and initiation, elongation and termination factors of translation.

7Frequent and clinically recognisable genetic causes of human diseases due to impaired mitochondrial translation are caused by mutations in mitochondrial tRNA synthetase and tRNA modifying genes.

8The potential interaction between cytosolic and mitochondrial translation requires further investigations.

9Mitochondrial diseases affect at least 1 in 5000 of the population and produce diverse clinical phenotypes often presented as multi-systemic disorders. In addition to the nucleus, human cells also harbour DNA in the mitochondria (mtDNA), which is essential for cell viability. This small (16.5 kb) genome is found in multiple copies in mitochondria, the subcellular organelles that often constitute more than 20% of the total cell volume. OXPHOS (oxidative phosphorylation) is responsible for the production of ATP by generating a proton gradient across the inner membrane of the mitochondria which is used by the mammalian cells. The mitochondrial OXPHOS system comprises around 150 different proteins out of which only 13 polypeptide subunits are encoded by the mtDNA. In addition, the mtDNA encodes the small and large rRNAs, and 22 distinct mitochondrial tRNAs that are necessary for the translation of only the mitochondrial-encoded proteins. The nuclear-encoded subunits of the respiratory chain (RC) complexes as well as proteins that are inevitable for normal mitochondrial protein synthesis (such as OXPHOS assembly, mtDNA metabolism and maintenance, mitochondrial cofactor biosynthesis, mitoribosomal subunits and assembly factors, regulators of mitochondrial expression and translation, etc.) are encoded by the nuclear genome (nDNA) and synthetised in the cytosol before transported into the organelle. The mitochondrial ribosomal proteins assemble with mitochondrial ribosomes 12S rRNA and 16S rRNA to form the mitochondrial ribosome. Lately it was also reported that import of 5S rRNA is also transported to the mitochondria being an essential component of the mitochondrial ribosomes.

10The components responsible for the proper mitochondrial translation are different from their cytosolic counterparts and they are more related to those of bacteria however the mechanisms of the translation follows the same major steps: initiation, elongation, termination and recycling of the ribosome (Fig. 1).

11The process of mitochondrial translation starts with the formation of the initiation complex. The separation of the two mitochondrial ribosomal subunits (28S and 39S) allow this complex to be formed which consists of the 28S subunit, mRNA and fMET-tRNA and IF2/3mt. This is followed by the entrance of the mRNA into the IF3mt:28S subunit complex. IF3mt is thought to support the correct position of mRNA to bind the small subunit at the peptidyl (P) site of the mitoribosome. When the appropriate start codon is present, the formylmethionin-tRNA can bind to the first codon with the help of IF2mt. The association of the mitoribosome stimulates the release of the initiation factors and the elongation on the 55S ribosome commence. MTFMT is critical for efficient human mitochondrial translation and reveal a human disorder of Met-tRNA(Met) formylation.

12To coordinate accurate and specific codon anti-codon pairing, mitochondrial elongation factor (EF-Tumt-GTP) and an amino-acylated tRNA arrives to the A-site of the mitoribosome. Upon correct codon-anticodon pairing the EF-Tumt-GDP leave the mitoribosome and the aminoacyl-tRNA moves into the P site where peptide bond formation is catalysed extending the growing polypeptide chain. EF-Tsmt plays a role as a nucleotide exchange factor and converts EF-Tumt to an active form (EF-Tumt-GTP). The GTP bound EFG1mt catalyses the translocation of the ribosome during the A and P site tRNAs move to the P and exit (E) sites of the mitoribosome. This elongation step repeats itself until the stop codon (UAA, UAG, AGA or AGG) is encountered in the A-site. Mutation of the mitochondrial elongation factors typically associated with encephalopathy and other organ involvement (liver, heart). Clinical symptoms are present in early infancy and the affected children die early.

13The mitochondrial release factor (mtRF1a) recognises the stop codon and binds to the mitoribosome, induces hydrolysis of the peptidyl-tRNA bond in the A-site releasing the mature protein from the site. Other termination release factors such as mtRF1, C12orf65 and ICT1 are also thought to play an essential role in the termination. As a last step mitochondrial recycling factors (mtRRF1 and mtRRF2) translocate to the A-site to induce the release of the mRNA. Up to date only the C12orf65 has been identified as a disease causing gene. Affected patients develop optic atrophy and ophthalmoplegia with Leigh syndrome.

14The expression of mitochondrial proteins is regulated by their own translational activators that bind mitochondrial mRNAs usually to their 5‘-untranslated regions, and each mitochondrial mRNA has its own translational activator(s), which has been first shown in yeast. Recent studies showed that these translational activators can be part of a feedback control loops which only permit translation if the downstream assembly of nascent translation products can occur. Recently mutations in nuclear-encoded translational activators of mitochondrial proteins such as TACO1 were also implicated in human disease. A regulatory role of aminoacyl-tRNA synthetases has been suggested in both cytosolic and mitochondrial translation and other factors, such as MTERF3 has been implicated to coordinate crosstalk between transcription and translation for the regulation of mammalian mtDNA.

15Impaired mitochondrial translation usually results in severe combined respiratory chain dysfunction through deficient function of all mtDNA-encoded proteins however some nuclear genes have been shown to alter the translation of single mitochondrial-encoded proteins. The defective mitochondrial proteins lead to deficient ATP production, and cellular energy deficit.

16Human cells contain 17 cytoplasmic ARS polypeptides, including the bifunctional glutamyl-prolyl-tRNA synthetase (EPRS), and 18 mitochondrial ARS2 enzymes. Three ARS genes encode proteins with dual localisation, present both in the cytoplasm and mitochondria (GARS, KARS, QARS), and the transport of the mitochondrial isoforms is ensured by a mitochondrial targeting signal.

17Beside disorders due to impaired mitochondrial translation, several human disorders are caused by altered cytosolic translation. Interestingly these diseases also lead to tissue specific clinical presentations mainly affecting brain, spinal cord and peripheral neurons, illustrated by clinical presentations such as Charcot–Marie–Tooth disease (CMT), distal hereditary motor neuropathies (dHMN) or leukoencephalopathy with vanishing white matter (VWM). Further implications of altered translation are highlighted by variable and complex clinical presentations, including diseases of eye, cartilage, skin, hair and even cancer (Table 1).

18As it was predicted, mutations in the nuclear genes coding for various components of the translation machinery could give rise to a wide spectrum of diseases and phenotypes (Table 1). Mitochondrial protein synthesis requires several nuclear-encoded factors, such as ribosomal proteins, ribosome assembly proteins, aminoacyl-tRNA synthetases, tRNA modifying enzymes and initiation, elongation and termination factors of translation (Fig. 1). Autosomal recessive mutations have been reported in several of these factors in association with variable clinical presentations. Here we note, that disorders of mitochondrial protein synthesis usually result in combined RC deficiencies and associated with abnormal mitochondria (ragged red fibres, COX negative fibres) on histology. However a defect of translation activation factors or post-transcriptional regulators of mammalian mtDNA expression may cause impairment in the stability of certain mitochondrial transcripts, as reported in patients with TACO1 and LRPPRC deficiency, respectively. Because these defects appear to affect only a single OXPHOS enzyme (COX), these patients show isolated COX deficiency.

19Frequent causes of impaired mitochondrial translation are mtDNA rearrangements (e.g. Kearns–Sayre syndrome) that affect mitochondrial tRNA and/or rRNA genes or single mt-tRNA point mutations. About half of the mtDNA mutations causing diseases in humans occur in tRNA genes (MELAS, MERRF, etc.) and the heterogeneous clinical manifestations usually reflect variable heteroplasmy. Homoplasmic tRNA mutations with variable penetrance and clinical presentations also occur and suggest the role of genetic or epigenetic modifiers in mitochondrial translation. Although most of these conditions are progressive and fatal, reversible infantile cytochrome c oxidase deficiency myopathy (or reversible infantile respiratory chain deficiency), due to a homoplasmic mt-tRNAGlu mutation stands out by showing spontaneous recovery.

20The currently defined disorders caused by nuclear defects of mitochondrial protein synthesis are usually early-onset, severe, often fatal diseases (Table 1) with extremely variable clinical presentations, and the reason behind is still unclear. Patients with translation elongation factor or mitochondrial ribosomal protein defects had an early age of onset and a severe multisystem disease with symptoms already present at birth or even prenatal in a few cases (Table 1).

21The extreme variability and relative strict tissue specificity of the diseases caused by mutations in mitochondrial tRNA synthetase genes illustrate the importance of understanding the factors influencing mitochondrial translation in different tissues.

22Some genes are selectively important in specific neuronal populations, as exemplified by leukoencephalopathy with brainstem and spinal cord involvement (LBSL) due to mutations in the mitochondrial aspartyl-tRNA synthetase 2 (DARS2), or pontocerebellar hypoplasia caused by argynyl tRNA synthetase 2 (RARS2) defect. Mutations in the glutamyl-tRNA synthetase (EARS2) cause early onset severe neurological disease (leukoencephalopathy involving the thalamus and brainstem with high lactate, LTBL). MTFMT deficiency leads to (relatively mild) Leigh syndrome with or without optic atrophy.

23Recently autosomal recessive mutations were reported in the AARS2 and MTO1 genes in patients with infantile hypertrophic cardiomyopathy. MTO1 encodes the enzyme that catalyzes the 5-carboxymethylamino-methylation of the wobble position in mt-tRNAGlu, mt-tRNAGln and mt-tRNALys. Patents with clinically indistinguishable clinical presentation of fatal infantile hypertrophic cardiomyopathy had mutations in the mitochondrial alanyl-tRNA synthetase 2 (AARS2) gene.

24Autosomal recessive mutations in the tRNA 5-methylaminomethyl-2-thiouridylate methyltransferase (TRMU), which is responsible for 2-thiouridylation of the mt-tRNAGlu, mt-tRNAGln and mt-tRNALys cause a severe but reversible infantile hepatopathy. Infants with reversible hepatopathy develop symptoms between 2 and 4 months of age, but if they survive this phase of liver failure, they recover and develop normally. The disease course and age of manifestation in TRMU deficiency shows remarkable similarities to reversible infantile myopathy and recent studies suggest that infantile cysteine concentrations may be important for the reversibility of both of these diseases.

25As a further complication of mt-tRNA synthetase dysfunction, the involvement of blood cells has been implicated by mutations in PUS1, resulting in mitochondrial myopathy, lactic acidosis and sideroblastic anaemia (MLASA). Mitochondrial tyrosyl-tRNA synthetase 2 (YARS2) mutations have been also identified in families with MLASA, very similar, but earlier onset compared to the phenotype caused by deficiency of a mt-tRNA modifying enzyme PUS1.

26Other characteristic, rare diseases are HyperUricemia, Pulmonary hypertension, Renal failure and Alkalosis (HUPRA) syndrome, which is caused by mutations in the mitochondrial seryl-tRNA synthetase 2 (SARS2) and Perrault syndrome, characterised by ovarian dysgenesis and sensorineural hearing loss due to mutations in the mitochondrial histidyl-tRNA synthetase 2 (HARS2) and leucyl-tRNA synthetase 2 (LARS2).

27Despite the rapid advances in technologies and the growing number of human disease genes and studies on mechanisms of mammalian mitochondrial translation, its regulation still remains largely unexplored. The variety and intriguing tissue and cell-type specific clinical presentations in both mitochondrial and cytosolic translation, and the dual function of some ARS enzymes suggest substantial interaction and overlap between these two protein synthesis pathways, which have not been extensively studied to date. Due to the abundant proteins and factors required for maintaining accurate mitochondrial translation, it is a challenge to identify the genetic defect in all cases. However rapid development of genetic technologies (next generation sequencing) resulted in a dynamic improvement in genetic diagnosis. Although there is a phenotypic diversity in patients with mitochondrial translation deficiencies, we observed some emerging clinical subgroups, which recently turned out to be associated with specific genetic defects. In patients with neurological presentation tRNA synthetases or tRNA modifying factors are the most likely cause of disease. AARS2 and MTO1 mutations are preferentially associated with cardiomyopathy, mutations in TRMU present with infantile, reversible liver failure, YARS2 and PUS1 mutations lead to sideroblastic anaemia and myopathy, and RMND1 deficiency cause deafness, myopathy, renal involvement and a severe biochemical defect (Table 1).

28Defining the exact pathomechanisms will suggest new avenues for treatment in these disorders, as it has been recently studied in reversible COX deficiency myopathy and TRMU deficiency. Downregulation of TRMU that is required for 2-thiouriylation in cells from patient with RIRCD led to a reduction in levels of mt-tRNAGlu thiolation resulting in a defect of mitochondrial protein synthesis. Cysteine is essential for normal thiolation and supplementation of L-cysteine improved mitochondrial gene translation not only in TRMU but also in RIRCD cells. Interesting experimental progresses are being pursued towards gene therapeutic approaches for mitochondrial translational disorders. Engineered human mitochondrial tRNAs and mRNAs – containing RP import sequence – can be efficiently imported into the mitochondria where they restore translation. The use of exosomes the body's own vehicle mechanism for delivering protein and genetic biomarkers are also promising and new avenues are being identified. Recently it has been shown that mRNAs for most tRNA synthetases can be detected in exosomes. The detection of mutations in factors involved in mitochondrial translation widens our understanding of mitochondrial disease and highlights basic molecular mechanisms.