bims-cytox1 2021-03-07 papers

Issue of 2021–03–07
five papers selected by
Gavin McStay, Staffordshire University

Vavilovskii Zhurnal Genet Selektsii. 2020 Aug;24(5): 512-518

Study of the effect of the introduction of mitochondrial import determinants into the gRNA structure on the activity of the gRNA/SpCas9 complex in vitro.

E G Zakirova, Y V Vyatkin, N A Verechshagina, V V Muzyka, I O Mazunin, K E Orishchenko.

It has long been known that defects in the structure of the mitochondrial genome can cause various neuromuscular and neurodegenerative diseases. Nevertheless, at present there is no effective method for treating mitochondrial diseases. The major problem with the treatment of such diseases is associated with mitochondrial DNA (mtDNA) heteroplasmy. It means that due to a high copy number of the mitochondrial genome, mutant copies of mtDNA coexist with wild-type molecules in the same organelle. The clinical symptoms of mitochondrial diseases and the degree of their manifestation directly depend on the number of mutant mtDNA molecules in the cell. The possible way to reduce adverse effects of the mutation is by shifting the level of heteroplasmy towards the wild-type mtDNA molecules. Using this idea, several gene therapeutic approaches based on TALE and ZF nucleases have been developed for this purpose. However, the construction of protein domains of such systems is rather long and laborious process. Meanwhile, the CRISPR/Cas9 system is fundamentally different from protein systems in that it is easy to use, highly efficiency and has a different mechanism of action. All the characteristics and capabilities of the CRISPR/Cas9 system make it a promising tool in mitochondrial genetic engineering. In this article, we demonstrate for the first time that the modification of gRNA by integration of specific mitochondrial import determinants in the gRNA scaffold does not affect the activity of the gRNA/Cas9 complex in vitro.

Keywords: CRISPR/Cas9; heteroplasmy; mitochondrial DNA; the mitochondrial import determinants

DOI: https://doi.org/10.18699/VJ20.643

Mol Diagn Ther. 2021 Mar 01.

Current and Emerging Clinical Treatment in Mitochondrial Disease.

Rory J Tinker, Albert Z Lim, Renae J Stefanetti, Robert McFarland.

Primary mitochondrial disease (PMD) is a group of complex genetic disorders that arise due to pathogenic variants in nuclear or mitochondrial genomes. Although PMD is one of the most prevalent inborn errors of metabolism, it often exhibits marked phenotypic variation and can therefore be difficult to recognise. Current treatment for PMD revolves around supportive and preventive approaches, with few disease-specific therapies available. However, over the last decade there has been considerable progress in our understanding of both the genetics and pathophysiology of PMD. This has resulted in the development of a plethora of new pharmacological and non-pharmacological therapies at varying stages of development. Many of these therapies are currently undergoing clinical trials. This review summarises the latest emerging therapies that may become mainstream treatment in the coming years. It is distinct from other recent reviews in the field by comprehensively addressing both pharmacological non-pharmacological therapy from both a bench and a bedside perspective. We highlight the current and developing therapeutic landscape in novel pharmacological treatment, dietary supplementation, exercise training, device use, mitochondrial donation, tissue replacement gene therapy, hypoxic therapy and mitochondrial base editing.

DOI: https://doi.org/10.1007/s40291-020-00510-6

J Mol Biol. 2021 Feb 24. pii: S0022-2836(21)00088-7. [Epub ahead of print] 166894

Building better barrels - β-barrel biogenesis and insertion in bacteria and mitochondria.

Kathryn A Diederichs, Susan K Buchanan, Istvan Botos.

β-barrel proteins are folded and inserted into outer membranes by multi-subunit protein complexes that are conserved across different types of outer membranes. In Gram-negative bacteria this complex is the barrel-assembly machinery (BAM), in mitochondria it is the sorting and assembly machinery (SAM) complex, and in chloroplasts it is the outer envelope protein Oep80. Mitochondrial β-barrel precursor proteins are translocated from the cytoplasm to the intermembrane space by the translocase of the outer membrane (TOM) complex, and stabilized by molecular chaperones before interaction with the assembly machinery. Outer membrane bacterial BamA interacts with four periplasmic accessory proteins, whereas mitochondrial Sam50 interacts with two cytoplasmic accessory proteins. Despite these major architectural differences between BAM and SAM complexes, their core proteins, BamA and Sam50, seem to function the same way. Based on the new SAM complex structures, we propose that the mitochondrial β-barrel folding mechanism follows the budding model with barrel-switching aiding in the release of new barrels. We also built a new molecular model for Tom22 interacting with Sam37 to identify regions that could mediate TOM-SAM supercomplex formation.

Keywords: BAM complex; Oep80; SAM complex; TOM complex; outer membrane beta-barrels

DOI: https://doi.org/10.1016/j.jmb.2021.166894

FEBS Lett. 2021 Mar 02.

Genetic basis of mitochondrial diseases.

Mirjana Gusic, Holger Prokisch.

Mitochondrial disorders are monogenic disorders characterized by a defect in oxidative phosphorylation and caused by pathogenic variants in one of over 340 different genes. The implementation of whole exome sequencing has led to a revolution in their diagnosis, duplicated the number of associated disease genes, and significantly increased the diagnosed fraction. However, the genetic etiology of a substantial fraction of patients exhibiting mitochondrial disorders remains unknown, highlighting limitations in variant detection and interpretation, which calls for improved computational and DNA sequencing methods, as well as the addition of OMICS tools. More intriguingly, this also suggests that some pathogenic variants lie outside of the protein-coding genes and that the mechanisms beyond the Mendelian inheritance and the mtDNA are of relevance. This review covers the current status of the genetic basis of mitochondrial diseases, discusses current challenges and perspectives, and explores the contribution of factors beyond the protein-coding regions and monogenic inheritance in the expansion of the genetic spectrum of disease.

Keywords: Diagnostics; Genetics; Metabolic disorders; Mitochondrial disease; Multi-omics; Non-coding; Oxidative phosphorylation; Variants

DOI: https://doi.org/10.1002/1873-3468.14068

Genes (Basel). 2021 Feb 20. pii: 300. [Epub ahead of print]12(2):

The Power of Yeast in Modelling Human Nuclear Mutations Associated with Mitochondrial Diseases.

Camilla Ceccatelli Berti, Giulia di Punzio, Cristina Dallabona, Enrico Baruffini, Paola Goffrini, Tiziana Lodi, Claudia Donnini.

The increasing application of next generation sequencing approaches to the analysis of human exome and whole genome data has enabled the identification of novel variants and new genes involved in mitochondrial diseases. The ability of surviving in the absence of oxidative phosphorylation (OXPHOS) and mitochondrial genome makes the yeast Saccharomyces cerevisiae an excellent model system for investigating the role of these new variants in mitochondrial-related conditions and dissecting the molecular mechanisms associated with these diseases. The aim of this review was to highlight the main advantages offered by this model for the study of mitochondrial diseases, from the validation and characterisation of novel mutations to the dissection of the role played by genes in mitochondrial functionality and the discovery of potential therapeutic molecules. The review also provides a summary of the main contributions to the understanding of mitochondrial diseases emerged from the study of this simple eukaryotic organism.

Keywords: diseases; mitochondria; yeast model

DOI: https://doi.org/10.3390/genes12020300