Biology subjects

Thoma, F.

Publications and source records attributed to Thoma, F..

4 recordsLinked to original sources

bioRxiv2026

RNA-dependent association of the pyruvate dehydrogenase complex with mtDNA-containing assemblies supports mitochondrial translation

Mitochondrial DNA (mtDNA) encodes core subunits of the oxidative phosphorylation machinery, and its expression is spatially organized, with the genome packaged into nucleoids around which transcription and mitoribosome assembly are concentrated. How this architecture is built, and how it is linked to the metabolic state of the organelle, remains poorly understood. Here we show that the pyruvate dehydrogenase complex (PDHc), which supplies acetyl-CoA to the tricarboxylic acid cycle, is a component of this machinery in Saccharomyces cerevisiae. PDHc co-purifies with mtDNA and resides in high-molecular-weight assemblies whose integrity requires RNA rather than DNA, and proximity labeling places it selectively adjacent to the mitoribosome. The E1 subunit Pda1 concentrates into discrete foci that depend on mtDNA and disperse reversibly upon inhibition of mitochondrial translation. Loss of Pda1, Pdb1 or Lat1, but not of the E3-binding protein Pdx1, compromises the maintenance of mtDNA; deletion of PDA1, PDB1 or LAT1 also reduces output from a mitochondrially encoded reporter, and PDHc-deficient cells are hypersensitive to translational inhibition. Catalytically inactive Pda1 and Lat1 variants rescue both defects as effectively as the wild-type proteins, demonstrating a function genetically separable from acetyl-CoA synthesis. PDHc therefore acts as a non-catalytic component of the RNA-dependent architecture that supports mitochondrial gene expression.

cell biology↗

bioRxiv2025

Local Mitochondrial Physiology Defined by mtDNA Quality Guides Purifying Selection

The mitochondrial genome (mtDNA) encodes essential subunits of the electron transport chain and ATP synthase. Mutations in these genes impair oxidative phosphorylation, compromise mitochondrial ATP production and cellular energy supply, and can cause mitochondrial diseases. These consequences highlight the importance of mtDNA quality control (mtDNA-QC), the process by which cells selectively maintain intact mtDNA to preserve respiratory function. Here, we developed a high-throughput flow cytometry assay for Saccharomyces cerevisiae to track mtDNA segregation in cell populations derived from heteroplasmic zygotes, in which wild-type (WT) mtDNA is fluorescently labeled and mutant mtDNA remains unlabeled. Using this approach, we observe purifying selection against mtDNA lacking subunits of complex III (COB), complex IV (COX2) or the ATP synthase (ATP6), under fermentative conditions that do not require respiratory activity. By integrating cytometric data with growth assays and simulations, we show that the decline of {Delta}atp6 mtDNA reflects both intracellular selection for intact genomes and a proliferative disadvantage of cells lacking ATP synthase. In contrast, loss of {Delta}cob and {Delta}cox2 mtDNA is primarily driven by selective maintenance of functional genomes. In heteroplasmic cells containing both intact and mutant mtDNA, fluorescent reporters revealed local reductions in ATP levels and membrane potential ({Delta}{Psi}) near mutant genomes, indicating spatial heterogeneity in mitochondrial physiology that reflects local mtDNA quality. Disruption of the respiratory chain by deletion of nuclear-encoded subunits (RIP1, COX4) abolished these physiological gradients and impaired mtDNA-QC, suggesting that local bioenergetic differences are required for selective recognition. Together, our findings support a model in which yeast cells assess local respiratory function as a proxy for mtDNA integrity, enabling intracellular selection for functional mitochondrial genomes.

cell biology↗

bioRxiv2024

Delayed protein translocation protects mitochondria against toxic CAT-tailed proteins

Ribosome-associated quality control (RQC) protects cells against toxic effects of faulty polypeptides produced by stalled ribosomes. However, mitochondria are vulnerable to C-terminal alanyl-and-threonyl (CAT)-tailed proteins that are generated in this process and faulty nuclear-encoded mitochondrial proteins are handled by the recently discovered mitoRQC. Here, we performed a genome-wide screen in yeast to identify additional proteins involved in mitoRQC. We found that Pth2, a peptidyl-tRNA hydrolase in the mitochondrial outer membrane, influences aggregation of CAT-tailed proteins without majorly affecting the CAT-tailing process itself. Peptidyl-tRNA hydrolase activity is essential during this process, yet the activity of Pth2 can be substituted by another peptidyl-tRNA hydrolase, upon proper localization. Our data suggest that Pth2 acts through modulating protein translocation and that mitochondrial proteostasis network is relieved through increased access of CAT-tailed proteins to cytosolic chaperones. Other hits obtained in the screen show that, in general, delayed protein translocation protects mitochondria against toxic CAT-tailed proteins.

cell biology↗

bioRxiv2021

YeastMate: Neural network-assisted segmentation of mating and budding events in S. cerevisiae

Here, we introduce YeastMate, a user-friendly deep learning-based application for automated detection and segmentation of Saccharomyces cerevisiae cells and their mating and budding events in microscopy images. We build upon Mask R-CNN with a custom segmentation head for the subclassification of mother and daughter cells during lifecycle transitions. YeastMate can be used directly as a Python library or through a stand-alone GUI application and a Fiji plugin as easy to use frontends. The source code for YeastMate is freely available at https://github.com/hoerlteam/YeastMate under the MIT license. We offer packaged installers for our whole software stack for Windows, macOS and Linux. A detailed user guide is available at https://yeastmate.readthedocs.io.

bioinformatics↗