bims-mitran 2022-09-04 papers

Issue of 2022‒09‒04
five papers selected by
Andreas Kohler

Mol Cell. 2022 Aug 23. pii: S1097-2765(22)00764-X. [Epub ahead of print]

The human mitochondrial genome contains a second light strand promoter.

Benedict G Tan, Christian D Mutti, Yonghong Shi, Xie Xie, Xuefeng Zhu, Pedro Silva-Pinheiro, Katja E Menger, Héctor Díaz-Maldonado, Wei Wei, Thomas J Nicholls, Patrick F Chinnery, Michal Minczuk, Maria Falkenberg, Claes M Gustafsson.

The human mitochondrial genome must be replicated and expressed in a timely manner to maintain energy metabolism and supply cells with adequate levels of adenosine triphosphate. Central to this process is the idea that replication primers and gene products both arise via transcription from a single light strand promoter (LSP) such that primer formation can influence gene expression, with no consensus as to how this is regulated. Here, we report the discovery of a second light strand promoter (LSP2) in humans, with features characteristic of a bona fide mitochondrial promoter. We propose that the position of LSP2 on the mitochondrial genome allows replication and gene expression to be orchestrated from two distinct sites, which expands our long-held understanding of mitochondrial gene expression in humans.

Keywords: DdCBE; LSP2; POLRMT; light strand promoter; mitochondria; mitochondrial DNA; mitochondrial gene expression; mitochondrial promoter; mtDNA; transcription

DOI: https://doi.org/10.1016/j.molcel.2022.08.011

Nucleic Acids Res. 2022 Aug 30. pii: gkac720. [Epub ahead of print]

A deafness-associated mitochondrial DNA mutation caused pleiotropic effects on DNA replication and tRNA metabolism.

Feilong Meng, Zidong Jia, Jing Zheng, Yanchun Ji, Jing Wang, Yun Xiao, Yong Fu, Meng Wang, Feng Ling, Min-Xin Guan.

In this report, we investigated the molecular mechanism underlying a deafness-associated m.5783C > T mutation that affects the canonical C50-G63 base-pairing of TΨC stem of tRNACys and immediately adjacent to 5' end of light-strand origin of mitochondrial DNA (mtDNA) replication (OriL). Two dimensional agarose gel electrophoresis revealed marked decreases in the replication intermediates including ascending arm of Y-fork arcs spanning OriL in the mutant cybrids bearing m.5783C > T mutation. mtDNA replication alterations were further evidenced by decreased levels of PolγA, Twinkle and SSBP1, newly synthesized mtDNA and mtDNA contents in the mutant cybrids. The m.5783C > T mutation altered tRNACys structure and function, including decreased melting temperature, conformational changes, instability and deficient aminoacylation of mutated tRNACys. The m.5783C > T mutation impaired the 5' end processing efficiency of tRNACys precursors and reduced the levels of tRNACys and downstream tRNATyr. The aberrant tRNA metabolism impaired mitochondrial translation, which was especially pronounced effects in the polypeptides harboring higher numbers of cysteine and tyrosine codons. These alterations led to deficient oxidative phosphorylation including instability and reduced activities of the respiratory chain enzyme complexes I, III, IV and intact supercomplexes overall. Our findings highlight the impact of mitochondrial dysfunction on deafness arising from defects in mitochondrial DNA replication and tRNA metabolism.

DOI: https://doi.org/10.1093/nar/gkac720

Nat Commun. 2022 Aug 30. 13(1): 5100

Structural basis for shape-selective recognition and aminoacylation of a D-armless human mitochondrial tRNA.

Bernhard Kuhle, Marscha Hirschi, Lili K Doerfel, Gabriel C Lander, Paul Schimmel.

Human mitochondrial gene expression relies on the specific recognition and aminoacylation of mitochondrial tRNAs (mtRNAs) by nuclear-encoded mitochondrial aminoacyl-tRNA synthetases (mt-aaRSs). Despite their essential role in cellular energy homeostasis, strong mutation pressure and genetic drift have led to an unparalleled sequence erosion of animal mtRNAs. The structural and functional consequences of this erosion are not understood. Here, we present cryo-EM structures of the human mitochondrial seryl-tRNA synthetase (mSerRS) in complex with mtRNASer(GCU). These structures reveal a unique mechanism of substrate recognition and aminoacylation. The mtRNASer(GCU) is highly degenerated, having lost the entire D-arm, tertiary core, and stable L-shaped fold that define canonical tRNAs. Instead, mtRNASer(GCU) evolved unique structural innovations, including a radically altered T-arm topology that serves as critical identity determinant in an unusual shape-selective readout mechanism by mSerRS. Our results provide a molecular framework to understand the principles of mito-nuclear co-evolution and specialized mechanisms of tRNA recognition in mammalian mitochondrial gene expression.

DOI: https://doi.org/10.1038/s41467-022-32544-1

Sci Rep. 2022 Aug 31. 12(1): 14804

A novel splice variant of Elp3/Kat9 regulates mitochondrial tRNA modification and function.

Rachid Boutoual, Hyunsun Jo, Indra Heckenbach, Ritesh Tiwari, Herbert Kasler, Chad A Lerner, Samah Shah, Birgit Schilling, Vincenzo Calvanese, Matthew J Rardin, Morten Scheibye-Knudsen, Eric Verdin.

Post-translational modifications, such as lysine acetylation, regulate the activity of diverse proteins across many cellular compartments. Protein deacetylation in mitochondria is catalyzed by the enzymatic activity of the NAD+-dependent deacetylase sirtuin 3 (SIRT3), however it remains unclear whether corresponding mitochondrial acetyltransferases exist. We used a bioinformatics approach to search for mitochondrial proteins with an acetyltransferase catalytic domain, and identified a novel splice variant of ELP3 (mt-ELP3) of the elongator complex, which localizes to the mitochondrial matrix in mammalian cells. Unexpectedly, mt-ELP3 does not mediate mitochondrial protein acetylation but instead induces a post-transcriptional modification of mitochondrial-transfer RNAs (mt-tRNAs). Overexpression of mt-ELP3 leads to the protection of mt-tRNAs against the tRNA-specific RNase angiogenin, increases mitochondrial translation, and furthermore increases expression of OXPHOS complexes. This study thus identifies mt-ELP3 as a non-canonical mt-tRNA modifying enzyme.

DOI: https://doi.org/10.1038/s41598-022-18114-x

J Biol Chem. 2022 Aug 24. pii: S0021-9258(22)00863-8. [Epub ahead of print] 102420

Functional characterization of two variants of mitochondrial topoisomerase TOP1MT that impact regulation of the mitochondrial genome.

Iman Al Khatib, Jingti Deng, Andrew Symes, Marina Kerr, Hongliang Zhang, Sharyin Huang, Yves Pommier, Aneal Khan, Timothy E Shutt.

TOP1MT encodes a mitochondrial topoisomerase that is important for mtDNA regulation, and is involved in mitochondrial replication, transcription, and translation. Two variants predicted to affect TOP1MT function (V1 - R198C and V2 - V338L) were identified by exome sequencing of a newborn with hypertrophic cardiomyopathy. As no pathogenic TOP1MT variants had been confirmed previously, we characterized these variants for their ability to rescue several TOP1MT functions in knockout cells. Consistent with these TOP1MT variants contributing to the patient phenotype, our comprehensive characterization suggests that both variants had impaired activity. Critically, we determined neither variant was able to restore steady state levels of mitochondrial-encoded proteins, nor to rescue oxidative phosphorylation when re-expressed in TOP1MT knockout cells. However, we found the two variants behaved differently in some respects; while the V1 variant was more efficient in restoring transcript levels, the V2 variant showed better rescue of mtDNA copy number and replication. These findings suggest that the different TOP1MT variants affect distinct TOP1MT functions. Altogether, these findings begin to provide insight into the many roles that TOP1MT plays in the maintenance and expression of the mitochondrial genome, and how impairments in this important protein may lead to human pathology.

Keywords: TOP1MT; mitochondria; mtDNA; replication; transcription; translation

DOI: https://doi.org/10.1016/j.jbc.2022.102420