Biology subjects

Zatulovskiy, E.

Publications and source records attributed to Zatulovskiy, E..

9 recordsLinked to original sources

bioRxiv2026

Protein-driven colloid-osmotic pressure controls nuclear size, organization, and function

The size of the cell nucleus is tightly controlled and changes in nuclear size correlate with altered nuclear function during development, cell differentiation and senescence. How nuclear size is regulated and whether changes thereof are functionally relevant remains unclear. Here, we demonstrate that nuclear size is determined by the osmotic pressure exerted by proteins in yeast, human cells and frog egg extracts. The biophysical model we establish solves the long-standing question of how the nuclear-to-cytoplasmic ratio is regulated and maintained. Furthermore, altering protein-driven osmotic balance modulates the physical properties of the nucleus, with a direct effect on chromatin organization and gene expression: Nuclear enlargement causes the dissolution of heterochromatic structures and derepression of subtelomeres and transposons, while simultaneously down-regulating highly expressed housekeeping genes. Importantly, these global transcriptional patterns closely mimic the gene expression changes that occur as yeast, human, and drosophila cells enlarge. Importantly, artificially forcing nuclear compression is sufficient to reverse these size-associated expression changes. Together, our findings provide a quantitative, mechanistic explanation for the coupling between the size of the nucleus and the cell, and they establish nuclear size as a modulator of chromatin organization and gene expression.

cell biology↗

bioRxiv2025

Cell size modulates ferroptosis susceptibility

Size is a fundamental property of cells that influences many aspects of their physiology. This is because cell size sets the scale for all subcellular components and drives changes in the composition of the proteome. Given that large and small cells differ in their biochemical composition, we hypothesize that they should also differ in how they respond to signals and make decisions. Here, we investigated how cell size affects susceptibility to cell death. We found that large cells are more resistant to ferroptosis caused by system xc- inhibition. Ferroptosis is a type of cell death characterized by the iron-dependent accumulation of toxic lipid peroxides. This process is opposed by cysteine-dependent lipid peroxide detoxification mechanisms. We found that larger cells exhibit higher concentrations of the cysteine-containing metabolite glutathione and lower concentrations of membrane lipid peroxides. Mechanistically, this can be explained by the fact that larger cells had lower concentrations of an enzyme that enriches cellular membranes with peroxidation-prone polyunsaturated fatty acids, ACSL4, and increased concentrations of the iron-chelating protein ferritin, the glutathione-producing enzymes glutamate-cysteine ligase and glutathione synthetase, and the lysosomal protease cathepsin B, which can catabolize cysteine-rich extracellular proteins to produce additional cystine for fueling the synthesis of glutathione. Taken together, our results highlight the significant impact of cell size on cellular function and survival, revealing a size-dependent vulnerability to ferroptosis that could influence therapeutic strategies based on this cell death pathway.

cell biology↗

bioRxiv2023

The G1/S transition is promoted by Rb degradation via the E3 ligase UBR5

Mammalian cells make the decision to divide at the G1/S transition in response to diverse signals impinging on the retinoblastoma protein Rb, a cell cycle inhibitor and tumor suppressor. Rb is inhibited by two parallel pathways. In the canonical pathway, Cyclin D-Cdk4/6 kinase complexes phosphorylate and inactivate Rb. In the second, recently discovered pathway, Rbs concentration decreases during G1 to promote cells progressing through the G1/S transition. However, the mechanisms underlying this second pathway are unknown. Here, we found that Rbs concentration drop in G1 and recovery in S/G2 is controlled by phosphorylation-dependent protein degradation. In early G1 phase, un- and hypo-phosphorylated Rb is targeted by the E3 ligase UBR5. UBR5 knockout cells have higher Rb concentrations in early G1, exhibit a lower G1/S transition rate, and are more sensitive to Cdk4/6 inhibition. This last observation suggests that UBR5 inhibition can strengthen the efficacy of Cdk4/6 inhibitor-based cancer therapies.

cell biology↗

bioRxiv2022

The cell cycle inhibitor RB is diluted in G1 and contributes to controlling cell size in the mouse liver

Every type of cell in an animal maintains a specific size, which likely contributes to its ability to perform its physiological functions. While some cell size control mechanisms are beginning to be elucidated through studies of cultured cells, it is unclear if and how such mechanisms control cell size in an animal. For example, it was recently shown that RB, the retinoblastoma protein, was diluted by cell growth in G1 to promote size-dependence of the G1/S transition. However, it remains unclear to what extent the RB-dilution mechanism controls cell size in an animal. We therefore examined the contribution of RB-dilution to cell size control in the mouse liver. The RB-dilution model has two requirements. First, manipulations changing RB concentration drive corresponding changes in cell size, and second, the endogenous RB concentration decreases with cell size in G1. We found that both these requirements were met. Genetic perturbations decreasing RB protein concentrations through inducible shRNA expression or through liverspecific Rb1 knockout reduced hepatocyte size, while perturbations increasing RB protein concentrations in an Fah-/- mouse model increased hepatocyte size. Moreover, RB concentration decreased in larger G1 hepatocytes while the concentrations of the cell cycle activators Cyclin D1 and E2f1 remained relatively constant. Lastly, we tested how Rb1 manipulations affected G1/S cell size control in primary hepatocytes using live cell imaging. Loss of Rb1 weakened cell size control, i.e., reduced the inverse correlation between how much cells grew in G1 and how large they were at birth. Taken together, our results show that an RB- dilution mechanism contributes to cell size control in the mouse liver by linking cell growth to the G1/S transition.

cell biology↗

bioRxiv2022

Chromatin-modifying enzymes as modulators of nuclear size during lineage differentiation

The mechanism of nuclear size determination and alteration during normal lineage development and cancer pathologies which is not fully understood. As recently reported, chromatin modification can change nuclear morphology. Therefore, we screened a range of pharmacological chemical compounds that impact the activity of chromatin-modifying enzymes, in order to get a clue of the specific types of chromatin-modifying enzymes that remarkably effect nuclear size and shape. We found that interrupted activity of chromatin-modifying enzymes is associated with nuclear shape abnormalities. Furthermore, the activity of chromatin-modifying enzymes perturbs cell fate determination in cellular maintenance and lineage commitment. Our results indicated that chromatin-modifying enzyme regulates cell fate decision during lineage differentiation and is associate with nuclear size alteration. SignificanceHere we described for the first time the modulation of chromatin-modifying enzymes remarkably effects nuclear shape, and perturbs cell fate determination in cellular maintenance and lineage fate commitment in normal stem cells and leukemia cancer cells. We found that the irregularities of nuclear contour were highly related to pharmacological inhibition of chromatin-modifying enzyme activity. After manipulating a histone demethylase named GASC1, we found that upregulation of GASC1 impairs the differentiation of hESCs to terminally differentiated neural cells. Moreover, upregulation of GASC1 impairs the proliferation of leukemia cells due to cell cycle arrest in the G0/G1 phase and indicates misshapen nuclei. This study suggested that chromatin-modifying enzyme regulates the nuclear contour related cell fate decision also including cancer cell fate determination.

cancer biology↗

bioRxiv2021

Increasing cell size remodels the proteome and promotes senescence

Cell size is tightly controlled in healthy tissues, but it is poorly understood how variations in cell size affect cell physiology. To address this, we employed a high-accuracy mass spectrometry-based approach to measure how the proteome changes with cell size. Protein concentration changes are widespread, measurable in both asynchronous and G1-arrested cell populations, and predicted by subcellular localization, size-dependent changes in mRNA concentrations, and protein turnover. As proliferating cells grow larger, protein concentration changes typically associated with cellular senescence are increasingly pronounced. This suggests that large size is a cause rather than just a consequence of cell senescence. Consistent with this hypothesis, larger cells are prone to replicative, DNA damage-, and CDK4/6i-induced senescence. Size-dependent changes to the proteome, including those associated with senescence, are not observed when an increase in cell size is accompanied by a similar increase in ploidy. This shows that proteome composition is determined by the DNA-to-ploidy ratio rather than cell size per se and that polyploidization is an elegant method to generate large non-senescent cells as is commonly found in nature. Together, our findings show how cell size could impact many aspects of cell physiology through remodeling the proteome, thereby providing a rationale for cell size control and polyploidization.

cell biology↗

bioRxiv2019

Constitutive expression of a fluorescent protein reports the size of live human cells

Cell size is intimately related to cell physiology because it sets the geometric scale of organelles and biosynthesis. A number of methods exist to measure different aspects of size of individual cells, but each has significant drawbacks. Here, we present an alternative method to measure the size of single human cells using a nuclear localized fluorescent protein expressed from a constitutive promoter. We validate this method by comparing it to several established cell size measurement strategies, including flow cytometry optical scatter, total protein dyes, and quantitative phase microscopy. We directly compare our fluorescent protein measurement to the commonly used measurement of nuclear volume and show that our measurements are more robust and less dependent on image segmentation. We apply our method to examine how cell size impacts the cell division cycle, which reaffirms the importance of G1/S size control. Importantly, combining our size reporter with fluorescent labeling of a different protein in a different color channel allows measurement of concentration dynamics using simple widefield fluorescence imaging. Thus, we expect our method will be of use to other researchers interested in the topics of cell size control and, more broadly, how dynamically changing protein concentrations control cell fates.

cell biology↗

bioRxiv2018

Cell growth dilutes the cell cycle inhibitor Rb to trigger cell division

Cell size is fundamental to function in different cell types across the human body because it sets the scale of organelle structures, biosynthesis, and surface transport1,2. Tiny erythrocytes squeeze through capillaries to transport oxygen, while the million-fold larger oocyte divides without growth to form the ~100 cell pre-implantation embryo. Despite the vast size range across cell types, cells of a given type are typically uniform in size likely because cells are able to accurately couple cell growth to division3-6. While some genes whose disruption in mammalian cells affects cell size have been identified, the molecular mechanisms through which cell growth drives cell division have remained elusive7-12. Here, we show that cell growth acts to dilute the cell cycle inhibitor Rb to drive cell cycle progression from G1 to S phase in human cells. In contrast, other G1/S regulators remained at nearly constant concentration. Rb is a stable protein that is synthesized during S and G2 phases in an amount that is independent of cell size. Equal partitioning to daughter cells of chromatin bound Rb then ensures that all cells at birth inherit a similar amount of Rb protein. RB overexpression increased cell size in tissue culture and a mouse cancer model, while RB deletion decreased cell size and removed the inverse correlation between cell size at birth and the duration of G1 phase. Thus, Rb-dilution by cell growth in G1 provides a long-sought cell autonomous molecular mechanism for cell size homeostasis.

cell biology↗

bioRxiv2018

Cyclin D-Cdk4,6 drives cell cycle progression via the retinoblastoma protein’s C-terminal helix

The cyclin-dependent kinases Cdk4 and Cdk6 form complexes with D-type cyclins to drive cell proliferation. A well-known target of cyclin D-Cdk4,6 is the retinoblastoma protein, Rb, which inhibits cell cycle progression until its inactivation by phosphorylation. However, the role of cyclin D-Cdk4,6 phosphorylation of Rb in cell cycle progression is unclear because Rb can be phosphorylated by other cyclin-Cdk complexes and cyclin D-Cdk4,6 complexes have other targets that may drive cell division. Here, we show that cyclin D-Cdk4,6 docks one side of an alpha-helix in the C-terminus of Rb, which is not recognized by cyclins E, A, and B. This helix-based docking mechanism is shared by the p107 and p130 Rb-family members across metazoans. Mutation of the Rb C-terminal helix prevents phosphorylation, promotes G1 arrest, and enhances Rbs tumor suppressive function. Our work conclusively demonstrates that the cyclin D-Rb interaction drives cell division and defines a new class of cyclin-based docking mechanisms.

cell biology↗