Biology subjects

Schmoller, K. M.

Publications and source records attributed to Schmoller, K. M..

17 recordsLinked to original sources

bioRxiv2026

Retrograde signalling mediates cellular adaptation to mitochondrial DNA copy number alterations

Eukaryotic cells maintain multiple copies of the mitochondrial genome, which is essential for cellular metabolism. Accordingly, alterations in the mitochondrial DNA (mtDNA) copy number are associated with severe human diseases and ageing. However, the mechanisms through which cells regulate mtDNA copy number and the cellular consequences of altered copy number remain poorly understood. Here, using budding yeast as a model, we show that mtDNA copy number is determined by the amount of three limiting factors, Mip1, Abf2, and Rim1. By synthetically tuning the concentrations of only these three proteins, we can modulate mtDNA dosage inside the cell. This revealed that cells are surprisingly robust to mtDNA copy number alterations, with increased copy numbers even accelerating cell growth. Our findings suggest that this robustness is due to protein dosage compensation and independent regulation of mitochondrial morphology. Mechanistically, we identified a critical role of the retrograde signalling pathway for this adaptation. We show that signalling from mitochondria to the nucleus is upregulated in cells with higher mtDNA copy number, and disruption of this regulation diminishes their faster growth. Taken together, our work reveals regulatory principles that enable cells to adapt to mtDNA copy number alterations.

cell biology↗

bioRxiv2025

Conserved and Lineage-Specific Roles of KEA-Mediated Ion Homeostasis in Chlamydomonas

In Arabidopsis thaliana, seamless plastid gene expression and development depend on finely balanced ion homeostasis across the inner envelope (IE) membrane, maintained by the K/H antiporters AtKEA1/2. To assess whether these functions are retained across mono- and polyplastidic representatives of the green lineage, we studied CrKEA1, the sole IE KEA homolog in the unicellular alga Chlamydomonas reinhardtii. Using CRISPR/Cas9, we generated a Cr-kea1 knockout mutant that exhibits impaired photoautotrophic growth, chloroplast deformation, and photoinhibition. Transcriptomics revealed strong induction of ribosome biogenesis genes and reduced abundance of transcripts associated with cell and plastid division. Further RNA analyses confirmed defects in stromal rRNA maturation of Cr-kea1, paralleling observations from Arabidopsis kea1kea2 mutants. Expression of CrKEA1 in Arabidopsis rescued growth and rRNA maturation in At-kea1kea2, demonstrating functional continuity after the ancient divergence between the two lineages. Cross-species transcriptomic comparisons further revealed that IE KEA loss elicits both shared and species-specific transcriptional responses: PhANG repression was conserved between algae and plants, whereas activation of the chloroplast unfolded protein response (cpUPR) and reduced expression of genes tied to cell-cycle and plastid fission occurred only in Chlamydomonas. Single-cell time-lapse imaging confirmed that Cr-kea1 exhibits an increased frequency of aberrant cytokinesis, unequal division, and division failure. Our findings demonstrate that while IE KEA transporters fulfill conserved roles in maintaining the conditions for plastid gene expression, their integration into broader cellular networks has diverged between unicellular chlorophytes and embryophytes (land plants). This underscores a lineage-dependent tuning of plastid-nucleus communication shaped by organismal complexity and plastid number. One-sentence summaryDisruption of KEA-mediated chloroplast ion homeostasis in Chlamydomonas reinhardtii reveals conserved and lineage-specific control of plastid rRNA processing and cell cycle progression.

plant biology↗

bioRxiv2025

Division Asymmetry Drives Cell Size Variability in Budding Yeast

Cell size variability within proliferating populations reflects the interdependent regulation of cell growth and division as well as intrinsically stochastic effects. In budding yeast, the G1/S transition exerts strong size control in daughter cells, which manifests as the inverse correlation between how big a cell is when it is born and how much it grows in G1. However, mutations affecting this size control checkpoint only modestly influence population-wide size variability, often altering the coefficient of variation (CV) only by [~]10%. To resolve this paradox, we combine computational modeling and live-cell imaging to identify the principal determinants of cell size variability. Using an experimentally validated stochastic model of the yeast cell cycle, we perform parameter sensitivity analysis and find that division asymmetry between mothers and daughters is the dominant driver of CV, outweighing the effects of G1/S size control. Experimental measurements across genetic perturbations and growth conditions confirm a strong correlation between mother-daughter size asymmetry and population CV. These findings reconcile previous observations and show how asymmetric division operates in concert with G1/S size control to govern cell size heterogeneity.

cell biology↗

bioRxiv2025

A Fkh1/2 binding site array in the WHI5 promoter drives sub-scaling transcription

Cells typically regulate their size within a relatively tight range by coupling growth to the cell division cycle using a dedicated set of molecular mechanisms. In budding yeast, cells are born with a similar amount of the G1/S inhibitor protein Whi5 that is then diluted by growth throughout G1. As cells grow, Whi5 concentration decreases and cells become more likely to enter the cell cycle. Cells are born in G1 with similar amounts of Whi5 because of the size-independent (sub-scaling) expression of WHI5 mRNA during S/G2/M phases and the equal partitioning of Whi5 protein at division. While the latter is known to be achieved by association with chromatin before anaphase, the mechanism for the former is poorly understood. Through systematic mutations of the WHI5 promoter, we discovered that WHI5s core promoter region located -126 to -75 base pairs upstream of the start codon is responsible for sub-scaling expression. This sequence contains a repeating array of binding sites for the transcription factors Fkh1 and Fkh2. Mutation of any of these sites, deletion of either FKH1 or FKH2, or preventing Fkh1 or Fkh2 dimerization weakens the sub-scaling of WHI5 transcription. Taken together with structural predictions and a mathematical model of cooperative Fkh-DNA binding, we conclude that WHI5s sub-scaling transcription is regulated by a Fkh1/2 heteropolymer binding an array of sites in its core promoter.

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↗

bioRxiv2025

When mitochondria fall apart: Unbalanced mitochondrial segregation triggers loss of mtDNA in the absence of mitochondrial fusion

Mitochondrial biogenesis and inheritance must be carefully regulated alongside cell division to ensure proper mitochondrial function and cell survival. The dynamics of the mitochondrial network, including fusion and fission, play a crucial role in mitochondrial inheritance by facilitating the distribution and quality control of mitochondria. In budding yeast, simultaneous inhibition of both fusion and fission leads to loss of mitochondrial DNA (mtDNA) integrity, resulting in an increased frequency of petite cells. Loss of mitochondrial fusion alone results in the complete loss of mtDNA. While the loss of mtDNA in the absence of mitochondrial fusion has been known for almost 30 years, the reason remained unclear. Here, we investigate the consequences of impaired mitochondrial fusion through depletion of the mitofusin Fzo1. We follow the emerging phenotype by live-cell imaging and the analysis of more than thirty thousand single cells across their cell cycle. Fzo1 depletion causes rapid mitochondrial fragmentation and a reduction in mitochondrial membrane potential, followed by a progressive decline of mtDNA content and cellular growth rate over several cell divisions. During division, Fzo1-depleted daughters obtain an unusually large amount of mitochondria, leaving the mother with too little. This results in a strong disbalance of mitochondrial mass in the population. Additionally, Fzo1-depleted cells lose the ability to adjust mtDNA synthesis to compensate for a low mitochondrial content. The combined effects of unequal distribution and reduced synthesis drive rapid mtDNA loss. These results show how fusion defects lead to mtDNA loss and mitochondrial dysfunction, contributing to understanding diseases linked to fusion defects.

cell biology↗

bioRxiv2024

SpotMAX: a generalist framework for multi-dimensional automatic spot detection and quantification

The analysis of spot-like structures is a widespread task in microscopy-based cell biology. Existing solutions are typically specific to single applications and do not use multi-dimensional information from 5D datasets. Therefore, experimental scientists often resort to subjective manual annotation. Here, we present SpotMAX, a generalist AI-driven framework for automated spot detection and quantification. SpotMAX leverages the full scope of multi-dimensional datasets with an easy-to-use interface and an embedded framework for cell segmentation and tracking. SpotMAX outperforms state-of-the-art tools, and in some cases, even expert human annotators. We applied SpotMAX across diverse experimental questions, ranging from meiotic crossover events in C. elegans to mitochondrial DNA dynamics in S. cerevisiae and telomere length in mouse stem cells, leading to new biological insights. With its flexibility in integrating AI workflows, we anticipate that SpotMAX will become the standard for spot analysis in microscopy data. Source code: https://github.com/SchmollerLab/SpotMAX

cell biology↗

bioRxiv2024

Single-cell imaging reveals a key role of Bck2 in budding yeast cell size adaptation to nutrient challenges

Cell size is tightly controlled to optimize cell function and varies broadly depending on the organism, cell type, and environment. The budding yeast S. cerevisiae has been successfully used as a model to gain insights into eukaryotic cell size control. Multiple regulators of cell size in steady-state conditions have been identified, such as the G1/S transition activators Cln3 and Bck2 and the inhibitor Whi5. Individual deletions of these regulators result in populations with altered mean cell volumes. However, size homeostasis remains largely intact. Here, we show that although the roles of Bck2 and Cln3 for cell size regulation appear largely redundant in steady-state, a switch from fermentable to non-fermentable growth media reveals a unique role for Bck2 in cell size adaptation to changing nutrients. We use live-cell microscopy and machine learning-assisted image analysis to track single cells and their progeny through the nutrient switch. We find that after the switch, bck2{Delta} cells experience longer cell cycle arrests and more arrest-associated enlargement than wild-type, whi5{Delta} or cln3{Delta} cells, indicating that Bck2 becomes the critical G1/S activator in changing nutrients. Our work demonstrates that studying size regulation during nutrient shifts to mimic the dynamic environments of free-growing microorganisms can resolve apparent redundancies observed in steady-state size regulation.

cell biology↗

bioRxiv2024

The origin of septin ring size control in budding yeast

The size of organelles and cellular structures needs to be tightly controlled and adjusted to the overall cell size to ensure cell function. A prominent example of a self-assembly process forming a structure whose size scales with cell size is Cdc42-driven cell polarization and subsequent septin ring formation in Saccharomyces cerevisiae. Despite extensive research, the mechanisms that determine Cdc42 cluster and septin ring size are still unclear. Combining computational modeling, live-cell imaging and genetic manipulations, we show here that positive feedback in the polarization pathway, together with the amount of polarity proteins increasing with cell size, can account for the increase of the Cdc42 cluster area with cell size, and as a consequence also the scaling of the septin ring diameter. We demonstrate that in bni1{Delta} cells, which have a larger septin ring compared to wild-type cells but a similarly sized Cdc42-GTP cluster, disruption of F-actin cable assembly and polarization toward the bud site results in diffuse exocytosis, causing septin ring enlargement. Furthermore, we find that in cells with impaired negative feedback in the polarization pathway, the Cdc42-GTP cluster area expands compared to wild-type cells, while the septin ring diameter remains mostly unchanged. This decoupling can be partially explained by a high recruitment rate of septin via polarity factors. Taken together, we provide insights into the origin of septin ring size control, in particular the scaling with cell volume, by integrating new experimental findings and mechanistic models of budding yeast polarization.

cell biology↗

bioRxiv2024

A Multiparametric Analysis Reveals Differential Behavior of Spheroid Cultures on Distinct Ultra-Low Attachment Plates Types

Spheroids have become principal three-dimensional biological models to study cancer, developmental processes, and drug efficacy. For spheroid generation, ultra-low attachment plates are noteworthy due to their simplicity, compatibility with automation, and experimental and commercial accessibility. Nonetheless, it is unknown whether and to what degree the plate type impacts spheroid formation and biology. This study employed automated brightfield microscopy to systematically compare the size and eccentricity of spheroids formed in six different plate types using four distinct human cell lines, i.e., CCD-1137Sk fibroblasts, HaCaT keratinocytes, and MDA-MB-231 and HT-29 cancer cells. Results showed that all plate types exhibited similar sphe-roid-forming capabilities, and the gross patterns of growth or shrinkage during four days after seeding were comparable. Yet, size and eccentricity varied systematically among specific cell lines and plate types. A confocal wholemount analysis by a novel pipeline of AI-based 3D-image analysis procedures revealed changes in cell proliferation, cell number, nuclear volume, and keratino-cyte differentiation, which were accompanied by altered YAP1-signals. The findings show that the plate type may influence the outcome of experimental campaigns. It is advisable to scan different plate types for the optimal configuration for a specific investigation instead of using one standard plate for all kinds of applications.

cancer biology↗

bioRxiv2023

An mTOR/RNA pol I axis shapes chromatin architecture in response to fasting

Chromatin architecture is a fundamental mediator of genome function. Fasting is a major environmental cue across the animal kingdom. Yet, how it impacts on 3D genome organization is unknown. Here, we show that fasting induces a reversible and large-scale spatial reorganization of chromatin in C. elegans. This fasting-induced 3D genome reorganization requires inhibition of the nutrient-sensing mTOR pathway, a major regulator of ribosome biogenesis. Remarkably, loss of transcription by RNA Pol I, but not RNA Pol II nor Pol III, induces a similar 3D genome reorganization in fed animals, and prevents the restoration of the fed-state architecture upon restoring nutrients to fasted animals. Our work documents the first large-scale chromatin reorganization triggered by fasting and reveals that mTOR and RNA Pol I shape genome architecture in response to nutrients.

cell biology↗

bioRxiv2023

Decoupling of transcript and protein concentrations ensures budding yeast histone homeostasis in different nutrient conditions

To maintain protein homeostasis in changing nutrient environments, cells must precisely control the amount of their proteins, despite the accompanying changes in cell growth and biosynthetic capacity. As nutrients are major regulators of cell cycle length and progression, a particular challenge arises for the nutrient-dependent regulation of cell cycle genes, which are periodically expressed during the cell cycle. One important example are histones, which are needed at a constant histone-to-DNA stoichiometry. Here we show that budding yeast achieves histone homeostasis in different nutrients through a decoupling of transcript and protein abundance. We find that cells downregulate histone transcripts in poor nutrients to avoid toxic histone overexpression, but produce constant amounts of histone proteins through nutrient-specific regulation of translation efficiency. Our findings suggest that this allows cells to balance the need for rapid histone production under fast growth conditions with the tight regulation required to avoid toxic overexpression in poor nutrients.

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

Cell-ACDC: a user-friendly toolset embedding state-of-the-art neural networks for segmentation, tracking and cell cycle annotations of live-cell imaging data

Live-cell imaging is a powerful tool to study dynamic cellular processes on the level of single cells with quantitative detail. Microfluidics enables parallel high-throughput imaging, creating a downstream bottleneck at the stage of data analysis. Recent progress on deep learning image analysis dramatically improved cell segmentation and tracking. Nevertheless, manual data validation and correction is typically still required and broadly used tools spanning the complete range of live-cell imaging analysis, from cell segmentation to pedigree analysis and signal quantification, are still needed. Here, we present Cell-ACDC, a user-friendly graphical user-interface (GUI)-based framework written in Python, for segmentation, tracking and cell cycle annotation. We included two state-of-the-art and high-accuracy deep learning models for single-cell segmentation of yeast and mammalian cells implemented in the most used deep learning frameworks TensorFlow and PyTorch. Additionally, we developed and implemented a cell tracking method and embedded it into an intuitive, semi-automated workflow for label-free cell cycle annotation of single cells. The open-source and modularized nature of Cell-ACDC will enable simple and fast integration of new deep learning-based and traditional methods for cell segmentation or downstream image analysis. Source codehttps://github.com/SchmollerLab/Cell_ACDC

cell biology↗

bioRxiv2020

Transcription coordinates histone amounts and genome content

Biochemical reactions typically depend on the concentrations of the molecules involved, and cell survival therefore critically depends on the concentration of proteins. To maintain constant protein concentrations during cell growth, global mRNA and protein synthesis rates are tightly linked to cell volume. While such regulation is appropriate for most proteins, certain cellular structures do not scale with cell volume. The most striking example of this is the genomic DNA, which doubles during the cell cycle and increases with ploidy, but is independent of cell volume. Here, we show that the amount of histone proteins is coupled to the DNA content, even though mRNA and protein synthesis globally increase with cell volume. As a consequence, and in contrast to the global trend, histone concentrations (i.e. amounts per volume) decrease with cell volume but increase with ploidy. We find that this distinct coordination of histone homeostasis and genome content is already achieved at the transcript level, and is an intrinsic property of histone promoters that does not require direct feedback mechanisms. Mathematical modelling and histone promoter truncations reveal a simple and generalizable mechanism to control the cell volume- and ploidy-dependence of a given gene through the balance of the initiation and elongation rates.

molecular biology↗

bioRxiv2020

Size-independent mRNA synthesis and chromatin-based partitioning mechanisms generate and maintain constant amounts of protein per cell

Biosynthesis scales with cell size such that protein concentrations generally remain constant as cells grow. As an exception, synthesis of the cell-cycle inhibitor Whi5 sub-scales with cell size so that its concentration is lower in larger cells to promote cell-cycle entry. Here, we find that a transcriptional control uncouples Whi5 synthesis from cell size and, screening for similar genes, identify histones as the major class of sub-scaling transcripts besides WHI5. Histone synthesis is thereby matched to genome content rather than cell size. Such sub-scaling proteins are challenged by asymmetric cell division because proteins are typically partitioned in proportion to new-born cell volume. To avoid this fate, Whi5 uses chromatin-binding to partition similar protein amounts to each new-born cell regardless of cell size. Finally, disrupting both Whi5 synthesis and chromatin-based partitioning compromises G1 size control. Thus, specific transcriptional and partitioning mechanisms determine protein sub-scaling to control cell size.

cell biology↗

bioRxiv2020

Whi5 is diluted and protein synthesis does not dramatically increase in pre-Start G1

In their manuscript, Litsios et al.1 report a new model for how cell growth and biosynthetic activity control the G1/S transition in budding yeast. In essence, Litsios et al. claim that Start is driven by an increasing concentration of the G1 cyclin Cln3 due to a dramatic acceleration of protein synthesis in pre-Start G1 and not by the dilution of the cell cycle inhibitor Whi5. While we previously reported that Start was in part driven by cell growth during G1 diluting out the Start inhibitor Whi52, Litsios et al. report that Whi5 remains at constant concentration during G1, and changes in Whi5 concentration therefore do not contribute to Start. Since Litsios et al. directly contradict several key points of our own model of how cell growth triggers Start, we decided to investigate their claims and data. More specifically, we decided to investigate Litsios et al.s three major claims: O_LIWhi5 concentration remains constant during G1 C_LIO_LICln3 concentration strongly increases prior to Start C_LIO_LIGlobal protein synthesis rates increase by 2-3 fold prior to Start C_LI We investigated each of these three claims and found that the evidence presented by Litsios et al. does not support their claims due to inadequate analysis methods and flaws in their experiments.

molecular biology↗