Biology subjects

Osman, C.

Publications and source records attributed to Osman, C..

9 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↗

bioRxiv2026

The mitochondrial cohibitin complex facilitates the biogenesis of inner membrane proteins

The inner membrane of mitochondria contains many membrane-embedded carrier proteins of the SLC25 family to facilitate the exchange of metabolites between the cytosol and mitochondria. These carriers use a specific import route for their biogenesis that relies on the TIM22 complex as an inner membrane translocase. The molecular details of carrier biogenesis are not well understood. Using an improved, desthiobiotin-based proximity labeling approach called Destni in living yeast cells, we identified the mitochondrial protein Aim11 as an interactor of newly imported carrier proteins. Aim11 forms a 160 kDa complex together with Iai11, Gep7 and Mtc1 in the mitochondrial inner membrane that we named the comrade-of-prohibitin (cohibitin) complex owing to its genetic interaction with prohibitins. Deletion of Aim11 impairs the import of carrier proteins into the inner membrane and renders cells hypersensitive to carrier overexpression. Our data suggest that the cohibitin complex plays a quality control function that supports the TIM22-mediated insertion of carrier proteins into the inner membrane of mitochondria. SummaryKizmaz et al. identified Aim11 as a novel quality control factor that facilitates the insertion of carrier proteins into the inner membrane of mitochondria. Aim11 is part of the membrane-embedded cohibitin complex which cooperates with prohibitins in inner membrane protein biogenesis.

biochemistry↗

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↗

bioRxiv2025

Mrx6 binds the Lon protease Pim1 N-terminal domain to confer selective substrate specificity and regulate mtDNA copy number

Mitochondrial DNA (mtDNA) copy number regulation remains incompletely understood, despite its importance in cellular function. In Saccharomyces cerevisiae, Mrx6 belongs to the Pet20-domain-containing protein family, consisting of Mrx6, Pet20, and Sue1. Notably, absence of Mrx6 leads to increased mtDNA copy number. Here, we identify the C-terminus of Mrx6 as essential for its stability and interaction with the mitochondrial matrix protein Mam33. Deletion of Mam33 mimics the effect of Mrx6 loss, resulting in elevated mtDNA copy number. Bioinformatics, mutational analyses, and immunoprecipitation studies reveal that a subcomplex of Mam33 and Mrx6 trimers interacts with the substrate recognition domain of the conserved mitochondrial Lon protease Pim1 through a bipartite motif in the Pet20 domain of Mrx6. Loss of Mrx6, its paralog Pet20, Mam33, or mutations disrupting the interaction between Mrx6 and Pim1 stabilize key proteins required for mtDNA maintenance, the RNA polymerase Rpo41 and the HMG-box-containing protein Cim1. We propose that Mrx6, alongside Pet20 and Mam33, regulates mtDNA copy number by modulating substrate degradation through Pim1. Additionally, Mrx6 loss alters Cim1s function, preventing the detrimental effect on mtDNA maintenance observed upon Cim1 overexpression. The presence of three Pet20-domain proteins in yeast implies broader roles of Lon protease substrate recognition beyond mtDNA regulation.

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↗

bioRxiv2022

mtHsp70 converts mitochondrial proteostasis distress into impaired protein import

Functional mitochondria are essential for cell viability and depend on protein import from the cytosol. Impaired protein import initiates various well-characterized cellular programs that rescue or remove dysfunctional mitochondria. However, the molecular mechanism that underlies the initial reduction of protein import into defective mitochondria remained unknown. Here, we found that the redistribution of mtHsp70, mitochondrial chaperone that is involved in both protein import and protein folding, regulates the efficiency of protein import. During early mitochondrial stress, before rescue programs are initiated and membrane potential is affected, mtHsp70-dependent import was specifically impaired and association of mtHsp70 with the import complex reduced. Even under non-stress conditions, the majority of mtHsp70 is found in a substrate-bound state. We propose that the availability of free mtHsp70 limits protein import into mitochondria during stress.

biochemistry↗

bioRxiv2022

An improved MS2-MCP imaging system with minimal perturbation of mRNA stability

The MS2-MCP imaging system is widely used to study the mRNA spatial distribution in living cells. Here, we report that the MS2-MCP system may destabilize the tagged mRNA by targeting it to the nonsense-mediated mRNA decay pathway. We introduce an improved version, which has minimal perturbation of the mRNA stability.

cell biology↗

bioRxiv2021

Regulation with cell size ensures mitochondrial DNA homeostasis during cell growth

To maintain stable DNA concentrations, proliferating cells need to coordinate DNA replication with cell growth. For nuclear DNA, eukaryotic cells achieve this by coupling DNA replication to cell cycle progression, ensuring that DNA is doubled exactly once per cell cycle. By contrast, mitochondrial DNA replication is typically not strictly coupled to the cell cycle, leaving the open question of how cells maintain the correct amount of mitochondrial DNA during cell growth. Here, we show that in budding yeast, mitochondrial DNA copy number increases with cell volume, both in asynchronously cycling populations and during G1 arrest. Our findings suggest that cell-volume-dependent mitochondrial DNA maintenance is achieved through nuclear encoded limiting factors, including the mitochondrial DNA polymerase Mip1 and the packaging factor Abf2, whose amount increases in proportion to cell volume. By directly linking mitochondrial DNA maintenance to nuclear protein synthesis, and thus cell growth, constant mitochondrial DNA concentrations can be robustly maintained without a need for cell-cycle-dependent regulation.

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↗