Mitochondria are eukaryotic organelles that generate most of the energy in the cell by oxidative phosphorylation (OXPHOS). Each mitochondrion contains multiple copies of a closed circular ...double-stranded DNA genome (mtDNA). Human (mammalian) mtDNA encodes 13 essential subunits of the inner membrane complex responsible for OXPHOS. These mRNAs are translated by the mitochondrial protein synthesis machinery, which uses the 22 species of mitochondrial tRNAs (mt tRNAs) encoded by mtDNA. The unique structural features of mt tRNAs distinguish them from cytoplasmic tRNAs bearing the canonical cloverleaf structure. The genes encoding mt tRNAs are highly susceptible to point mutations, which are a primary cause of mitochondrial dysfunction and are associated with a wide range of pathologies. A large number of nuclear factors involved in the biogenesis and function of mt tRNAs have been identified and characterized, including processing endonucleases, tRNA-modifying enzymes, and aminoacyl-tRNA synthetases. These nuclear factors are also targets of pathogenic mutations linked to various diseases, indicating the functional importance of mt tRNAs for mitochondrial activity.
Mitochondria generate most cellular energy via oxidative phosphorylation. Twenty-two species of mitochondrial (mt-)tRNAs encoded in mtDNA translate essential subunits of the respiratory chain ...complexes. mt-tRNAs contain post-transcriptional modifications introduced by nuclear-encoded tRNA-modifying enzymes. They are required for deciphering genetic code accurately, as well as stabilizing tRNA. Loss of tRNA modifications frequently results in severe pathological consequences. Here, we perform a comprehensive analysis of post-transcriptional modifications of all human mt-tRNAs, including 14 previously-uncharacterized species. In total, we find 18 kinds of RNA modifications at 137 positions (8.7% in 1575 nucleobases) in 22 species of human mt-tRNAs. An up-to-date list of 34 genes responsible for mt-tRNA modifications are provided. We identify two genes required for queuosine (Q) formation in mt-tRNAs. Our results provide insight into the molecular mechanisms underlying the decoding system and could help to elucidate the molecular pathogenesis of human mitochondrial diseases caused by aberrant tRNA modifications.
In eukaryotes and archaea, tRNA genes frequently contain introns, which are removed during maturation. However, biological roles of tRNA introns remain elusive. Here, we constructed a complete set of ...Saccharomyces cerevisiae strains in which the introns were removed from all the synonymous genes encoding 10 different tRNA species. All the intronless strains were viable, but the tRNAPheGAA and tRNATyrGUA intronless strains displayed slow growth, cold sensitivity and defective growth under respiratory conditions, indicating physiological importance of certain tRNA introns. Northern analyses revealed that removal of the introns from genes encoding three tRNAs reduced the amounts of the corresponding mature tRNAs, while it did not affect aminoacylation. Unexpectedly, the tRNALeuCAA intronless strain showed reduced 5.8S rRNA levels and abnormal nucleolar morphology. Because pseudouridine (Ψ) occurs at position 34 of the tRNAIleUAU anticodon in an intron-dependent manner, tRNAIleUAU in the intronless strain lost Ψ34. However, in a portion of tRNAIleUAU population, position 34 was converted into 5-carbamoylmethyluridine (ncm5U), which could reduce decoding fidelity. In summary, our results demonstrate that, while introns are dispensable for cell viability, some introns have diverse roles, such as ensuring proper growth under various conditions and controlling the appropriate anticodon modifications for accurate pairing with the codon.
The steady-state levels of microRNAs (miRNAs) and their activities are regulated by the post-transcriptional processes. It is known that 3' ends of several miRNAs undergo post-dicing adenylation or ...uridylation. We isolated the liver-specific miR-122 from human hepatocytes and mouse livers. Direct analysis by mass spectrometry revealed that one variant of miR-122 has a 3'-terminal adenosine that is introduced after processing by Dicer. We identified GLD-2, which is a regulatory cytoplasmic poly(A) polymerase, as responsible for the 3'-terminal adenylation of miR-122 after unwinding of the miR-122/miR-122* duplex. In livers from GLD-2-null mice, the steady-state level of the mature form of miR-122 was specifically lower than in heterozygous mice, whereas no reduction of pre-miR-122 was observed, demonstrating that 3'-terminal adenylation by GLD-2 is required for the selective stabilization of miR-122 in the liver.
Mitochondrial RNAs are modified posttranscriptionally. These modifications are required for proper functioning of RNA molecules, and thereby contribute to essential mitochondrial processes. Herein, ...we describe our latest mass spectrometry-based platform for analysis of posttranscriptional modifications of mitochondrial tRNAs, and measuring the in vitro activity of mitochondrial RNA-modifying enzymes.
Mitochondrial DNA mutations that cause mitochondrial dysfunction are responsible for a wide spectrum of human diseases, referred to as mitochondrial diseases. Pathogenic point mutations are found ...frequently in genes encoding mitochondrial (mt) tRNAs, indicating that impaired functioning of mutant mt tRNAs is the primary cause of mitochondrial dysfunction. Our previous studies revealed the absence of posttranscriptional taurine modification at the anticodon wobble uridine in mutant mt tRNAs isolated from cells derived from patients with two major classes of mitochondrial diseases, MELAS (mitochondrial myopathy, encephalopathy, lactic acidosis, and stroke‐like episodes) and MERRF (myoclonus epilepsy associated with ragged red fibers). Defective taurine modification of the mutant mt tRNAs results in a deficiency in protein synthesis as the cognate codons of the mutant mt tRNA cannot be decoded. These findings represent the first evidence of a molecular pathogenesis caused by an RNA modification disorder. WIREs RNA 2011 2 376–386 DOI: 10.1002/wrna.65
This article is categorized under:
RNA-Based Catalysis > Miscellaneous RNA-Catalyzed Reactions
RNA Interactions with Proteins and Other Molecules > Protein–RNA Interactions: Functional Implications
RNA Processing > tRNA Processing
Ribosome biogenesis requires multiple assembly factors. InEscherichia coli, deletion of RlmE, the methyltransferase responsible for the 2′-O-methyluridine modification at position 2552 (Um2552) in ...helix 92 of the 23S rRNA, results in slow growth and accumulation of the 45S particle. We demonstrate that the 45S particle that accumulates in ΔrlmEis a genuine precursor that can be assembled into the 50S subunit. Indeed, 50S formation from the 45S precursor could be promoted by RlmE-mediated Um2552 formation in vitro. Ribosomal protein L36 (encoded byrpmJ) was completely absent from the 45S precursor in ΔrlmE, and we observed a strong genetic interaction betweenrlmEandrpmJ. Structural probing of 23S rRNA and high-salt stripping of 45S components revealed that RlmE-mediated methylation promotes interdomain interactions via the association between helices 92 and 71, stabilized by the single 2′-O-methylation of Um2552, in concert with the incorporation of L36, triggering late steps of 50S subunit assembly.
A subset of mitochondrial tRNAs (mt-tRNAs) contains taurine-derived modifications at 34U of the anticodon. Loss of taurine modification has been linked to the development of mitochondrial diseases, ...but the molecular mechanism is still unclear. Here, we showed that taurine modification is catalyzed by mitochondrial optimization 1 (Mto1) in mammals. Mto1 deficiency severely impaired mitochondrial translation and respiratory activity. Moreover, Mto1-deficient cells exhibited abnormal mitochondrial morphology owing to aberrant trafficking of nuclear DNA-encoded mitochondrial proteins, including Opa1. The mistargeted proteins were aggregated and misfolded in the cytoplasm, which induced cytotoxic unfolded protein response. Importantly, application of chemical chaperones successfully suppressed cytotoxicity by reducing protein misfolding and increasing functional mitochondrial proteins in Mto1-deficient cells and mice. Thus, our results demonstrate the essential role of taurine modification in mitochondrial translation and reveal an intrinsic protein homeostasis network between the mitochondria and cytosol, which has therapeutic potential for mitochondrial diseases.
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•Mto1-mediated taurine modification of mt-tRNAs is indispensable for protein translation•Mto1 deficiency induces mitochondrial protein imbalance and impairs membrane integrity•Mitochondrial proteins are aggregated in the cytosol and induce cytotoxic UPR•Chemical chaperones suppress proteostress and restore mitochondrial function via Opa1
Taurine modification of mitochondrial tRNA is associated with mitochondrial disease. Fakruddin et al. find that taurine modification is indispensable for mitochondrial protein translation. The authors also find that deficiency of taurine modification impairs a mitochondrial-cytosolic proteostatic network through an Opa1-dependent mechanism and demonstrate the therapeutic potential of chemical chaperones.
The imbalance between cholinergic activity and dopaminergic activity in the striatum causes a variety of neurological disorders, such as Parkinson's disease. During sensorimotor learning, the arrival ...of a conditioned stimulus reporting a reward evokes a pause response in the firing of the tonically active cholinergic interneurons in targeted areas of the striatum, whereas the same stimulus triggers an increase in the firing frequency of the dopaminergic neurons in the substantia nigra pars compacta. The pause response of the cholinergic interneurons begins with an initial depolarizing phase followed by a pause in spike firing and ensuing rebound excitation. The timing of the pause phase coincides well with the surge in dopaminergic firing, indicating that a dramatic rise in dopamine (DA) release occurs while nicotinic receptors remain unbound by acetylcholine. The pause response begins with dopamine D5 receptor‐dependent synaptic plasticity in the cholinergic neurons and an increased GABAergic IPSP, which is followed by a long pause in firing through D2 and D5 receptor‐dependent modulation of ion channels. Inactivation of muscarinic receptors on the projection neurons eventually yields endocannabinoid‐mediated, dopamine‐dependent long‐term depression in the medium spiny projection neurons. Breakdown of acetylcholine‐dopamine balance hampers proper functioning of the cortico‐basal ganglia‐thalamocortical loop circuits. In Parkinson's disease, dopamine depletion blocks autoinhibition of acetylcholine release through muscarinic autoreceptors, leading to excessive acetylcholine release which eventually prunes spines of the indirect‐pathway projection neurons of the striatum and thus interrupts information transfer from motor command centers in the cerebral cortex. Geriatr Gerontol Int 2010; 10 (Suppl. 1): S148–S157.
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