bims-mitran 2021-11-28 papers

Issue of 2021–11–28
four papers selected by
Andreas Kohler, Stockholm University

Proc Natl Acad Sci U S A. 2021 Nov 30. pii: e2111862118. [Epub ahead of print]118(48):

Structure of the translating Neurospora ribosome arrested by cycloheximide.

Lunda Shen, Zhaoming Su, Kailu Yang, Cheng Wu, Thomas Becker, Deborah Bell-Pedersen, Junjie Zhang, Matthew S Sachs.

Ribosomes translate RNA into proteins. The protein synthesis inhibitor cycloheximide (CHX) is widely used to inhibit eukaryotic ribosomes engaged in translation elongation. However, the lack of structural data for actively translating polyribosomes stalled by CHX leaves unanswered the question of which elongation step is inhibited. We elucidated CHX's mechanism of action based on the cryo-electron microscopy structure of actively translating Neurospora crassa ribosomes bound with CHX at 2.7-Å resolution. The ribosome structure from this filamentous fungus contains clearly resolved ribosomal protein eL28, like higher eukaryotes but unlike budding yeast, which lacks eL28. Despite some differences in overall structures, the ribosomes from Neurospora, yeast, and humans all contain a highly conserved CHX binding site. We also sequenced classic Neurospora CHX-resistant alleles. These mutations, including one at a residue not previously observed to affect CHX resistance in eukaryotes, were in the large subunit proteins uL15 and eL42 that are part of the CHX-binding pocket. In addition to A-site transfer RNA (tRNA), P-site tRNA, messenger RNA, and CHX that are associated with the translating N. crassa ribosome, spermidine is present near the CHX binding site close to the E site on the large subunit. The tRNAs in the peptidyl transferase center are in the A/A site and the P/P site. The nascent peptide is attached to the A-site tRNA and not to the P-site tRNA. The structural and functional data obtained show that CHX arrests the ribosome in the classical PRE translocation state and does not interfere with A-site reactivity.

Keywords: cryo-electron microscopy; protein synthesis; translation

DOI: https://doi.org/10.1073/pnas.2111862118

BMC Genom Data. 2021 Nov 26. 22(1): 52

The mtDNA mutation spectrum in the PolG mutator mouse reveals germline and somatic selection.

Kendra D Maclaine, Kevin A Stebbings, Daniel A Llano, Justin C Havird.

BACKGROUND: Mitochondrial DNA (mtDNA) codes for products necessary for electron transport and mitochondrial gene translation. mtDNA mutations can lead to human disease and influence organismal fitness. The PolG mutator mouse lacks mtDNA proofreading function and rapidly accumulates mtDNA mutations, making it a model for examining the causes and consequences of mitochondrial mutations. Premature aging in PolG mice and their physiology have been examined in depth, but the location, frequency, and diversity of their mtDNA mutations remain understudied. Identifying the locations and spectra of mtDNA mutations in PolG mice can shed light on how selection shapes mtDNA, both within and across organisms.
RESULTS: Here, we characterized somatic and germline mtDNA mutations in brain and liver tissue of PolG mice to quantify mutation count (number of unique mutations) and frequency (mutation prevalence). Overall, mtDNA mutation count and frequency were the lowest in the D-loop, where an mtDNA origin of replication is located, but otherwise uniform across the mitochondrial genome. Somatic mtDNA mutations have a higher mutation count than germline mutations. However, germline mutations maintain a higher frequency and were also more likely to be silent. Cytosine to thymine mutations characteristic of replication errors were the plurality of basepair changes, and missense C to T mutations primarily resulted in increased protein hydrophobicity. Unlike wild type mice, PolG mice do not appear to show strand asymmetry in mtDNA mutations. Indel mutations had a lower count and frequency than point mutations and tended to be short, frameshift deletions.
CONCLUSIONS: Our results provide strong evidence that purifying selection plays a major role in the mtDNA of PolG mice. Missense mutations were less likely to be passed down in the germline, and they were less likely to spread to high frequencies. The D-loop appears to have resistance to mutations, either through selection or as a by-product of replication processes. Missense mutations that decrease hydrophobicity also tend to be selected against, reflecting the membrane-bound nature of mtDNA-encoded proteins. The abundance of mutations from polymerase errors compared with reactive oxygen species (ROS) damage supports previous studies suggesting ROS plays a minimal role in exacerbating the PolG phenotype, but our findings on strand asymmetry provide discussion for the role of polymerase errors in wild type organisms. Our results provide further insight on how selection shapes mtDNA mutations and on the aging mechanisms in PolG mice.

Keywords: Germline mutations; Mutation spectrum; PolG; Protein hydrophobicity; ROS; mitochondrial theory of aging; mtDNA; mtDNA mutations; mtDNA selection

DOI: https://doi.org/10.1186/s12863-021-01005-x

Trends Cell Biol. 2021 Nov 23. pii: S0962-8924(21)00207-5. [Epub ahead of print]

Rewiring cell signalling pathways in pathogenic mtDNA mutations.

Chih-Yao Chung, Gabriel E Valdebenito, Anitta R Chacko, Michael R Duchen.

Mitochondria generate the energy to sustain cell viability and serve as a hub for cell signalling. Their own genome (mtDNA) encodes genes critical for oxidative phosphorylation. Mutations of mtDNA cause major disease and disability with a wide range of presentations and severity. We review here an emerging body of data suggesting that changes in cell metabolism and signalling pathways in response to the presence of mtDNA mutations play a key role in shaping disease presentation and progression. Understanding the impact of mtDNA mutations on cellular energy homeostasis and signalling pathways seems fundamental to identify novel therapeutic interventions with the potential to improve the prognosis for patients with primary mitochondrial disease.

Keywords: cell signalling; heteroplasmy; metabolic remodelling; mitochondrial disease; mtDNA

DOI: https://doi.org/10.1016/j.tcb.2021.10.005

Int J Mol Sci. 2021 Nov 12. pii: 12223. [Epub ahead of print]22(22):

A Yeast-Based Repurposing Approach for the Treatment of Mitochondrial DNA Depletion Syndromes Led to the Identification of Molecules Able to Modulate the dNTP Pool.

Giulia di Punzio, Micol Gilberti, Enrico Baruffini, Tiziana Lodi, Claudia Donnini, Cristina Dallabona.

Mitochondrial DNA depletion syndromes (MDS) are clinically heterogenous and often severe diseases, characterized by a reduction of the number of copies of mitochondrial DNA (mtDNA) in affected tissues. In the context of MDS, yeast has proved to be both an excellent model for the study of the mechanisms underlying mitochondrial pathologies and for the discovery of new therapies via high-throughput assays. Among the several genes involved in MDS, it has been shown that recessive mutations in MPV17 cause a hepatocerebral form of MDS and Navajo neurohepatopathy. MPV17 encodes a non selective channel in the inner mitochondrial membrane, but its physiological role and the nature of its cargo remains elusive. In this study we identify ten drugs active against MPV17 disorder, modelled in yeast using the homologous gene SYM1. All ten of the identified molecules cause a concomitant increase of both the mitochondrial deoxyribonucleoside triphosphate (mtdNTP) pool and mtDNA stability, which suggests that the reduced availability of DNA synthesis precursors is the cause for the mtDNA deletion and depletion associated with Sym1 deficiency. We finally evaluated the effect of these molecules on mtDNA stability in two other MDS yeast models, extending the potential use of these drugs to a wider range of MDS patients.

Keywords: MIP1; MPV17; POLG; RNR2; RRM2B; SYM1; drug repurposing; mitochondrial DNA depletion syndromes (MDS); mitochondrial dNTP pool; yeast

DOI: https://doi.org/10.3390/ijms222212223