Biology subjects

Dey, G.

Publications and source records attributed to Dey, G..

29 records · Page 2Linked to original sources

bioRxiv2023

Life cycle-coupled evolution of mitosis in close relatives of animals

Eukaryotes have evolved towards one of two extremes along a spectrum of strategies for remodelling the nuclear envelope (NE) during cell division: disassembling the NE in an open mitosis or constructing an intranuclear spindle in a closed mitosis. Both classes of mitotic remodelling involve key differences in the core division machinery, but the evolutionary reasons for adopting a specific mechanism are unclear. Here, we use an integrated comparative genomics and ultrastructural imaging approach to investigate mitotic strategies in Ichthyosporea, close relatives of animals and fungi. We show that species within this clade have diverged towards either a fungal-like closed or an animal-like open mitosis, most likely to support distinct multi- or uninucleated states. Our results suggest that multinucleated life cycles favour the evolution of closed mitosis. One-Sentence SummaryMitotic specialization in animal relatives reveal that multinucleated life cycles favor the evolution of closed mitosis

cell biology↗

bioRxiv2022

Ultrastructure Expansion Microscopy reveals the nanoscale cellular architecture of budding and fission yeast

The budding yeast Saccharomyces cerevisiae and the fission yeast Schizosaccharomyces pombe have served as invaluable model organisms to study various fundamental and highly conserved cellular processes. While super-resolution (SR) microscopy has in recent years paved the way to a better understanding of the spatial organization of molecules in cells, its wide use in yeast models has remained limited due to the specific know-how and specialized instrumentation required, contrasted with the relative ease of endogenous tagging and live cell fluorescence microscopy in these systems. To facilitate SR microscopy in yeasts, we have extended the ultrastructure expansion microscopy (U-ExM) method to both S. cerevisiae and S. pombe, enabling 4-fold isotropic expansion in both systems. We demonstrate here that U-ExM allows the nanoscale imaging of the microtubule cytoskeleton and its associated spindle pole body (SPB), notably unveiling a conserved Sfi1p/Cdc31p spatial organization on the appendage bridge structure. In S. pombe, we validate the method by quantifying the homeostatic regulation of nuclear pore complex (NPC) number through the cell cycle. Combined with pan-labelling (NHS ester), which provides a global cellular context, U-ExM unveils the subcellular organization of the eukaryote yeast models S. cerevisiae and S. pombe. This easy-to-implement imaging with conventional microscopes provides nanoscale resolution and adds a powerful new method to the already extensive yeast toolbox.

cell biology↗

bioRxiv2022

A quantitative and spatial analysis of cell cycle regulators during the fission yeast cell cycle

We have carried out a systems-level analysis of the spatial and temporal dynamics of cell cycle regulators in the fission yeast Schizosaccharomyces pombe. In a comprehensive single cell analysis we have precisely quantified the levels of 38 proteins previously identified as regulators of the G2 to mitosis transition, and of 7 proteins acting at the G1 to S-phase transition. Only two of the 38 mitotic regulators exhibit changes in concentration at the whole cell level, the mitotic B-type cyclin Cdc13 which accumulates continually throughout the cell cycle, and the regulatory phosphatase Cdc25 which exhibits a complex cell cycle pattern. Both proteins show similar patterns of change within the nucleus as in the whole cell but at higher concentrations. In addition, the concentrations of the major fission yeast cyclin dependent kinase (CDK) Cdc2, the CDK regulator Suc1 and the inhibitory kinase Wee1 also increase in the nucleus peaking at mitotic onset but are constant in the whole cell. The significant increase in concentration with size for Cdc13 supports the model that mitotic B-type cyclin accumulation acts as a cell size sensor. We propose a two-step process for the control of mitosis. First, Cdc13 accumulates in a size-dependent manner which drives increasing CDK activity. Second, from mid G2 the increasing nuclear accumulation of Cdc25 and the counteracting Wee1 introduces a bistability switch that results in a rapid rise of CDK activity at the end of G2 and thus brings about an orderly progression into mitosis. Significance StatementAcross eukaryotes the increasing level of cyclin dependent kinase (CDK) activity drives progression through the cell cycle. As most cells divide at specific sizes, information responding to the size of the cell must feed into the regulation of CDK activity. In this study, we use fission yeast to precisely measure how proteins that have been previously identified in genome wide screens as cell cycle regulators change in their levels with cell cycle progression. We identify the mitotic B-type cyclin Cdc13 and mitotic inhibitory phosphatase Cdc25 as the only two proteins that change in both whole cell and nuclear concentration through the cell cycle, making them candidates for universal cell size sensors at the onset of mitosis and cell division.

cell biology↗

bioRxiv2021

Lymphocyte-specific Protein Tyrosine Kinase Regulates Homologous Recombination (HR) DNA Repair and Targeting Enhances PARPi Utility in HR Proficient Ovarian Cancer

Poly-ADP Ribose Polymerase (PARP) targeted therapy is clinically approved for the treatment of homologous recombination (HR) repair deficient tumors. The remarkable success in treatment of HR repair deficient cancers has not translated to HR-proficient cancer. Our studies identify the mechanism of non-receptor lymphocyte-specific protein tyrosine kinase (LCK) in HR repair in endometrioid epithelial ovarian cancer (eEOC). LCK expression is induced and activation in the nucleus in response to DNA damage insult. LCK inhibition attenuates expression of RAD51, BRCA1, and BRCA2 proteins necessary for HR-mediated DNA repair, sufficient to suppress RAD51 foci formation, and augments {gamma}H2AX foci formation. Mechanistically, DNA damage leads to direct interaction of LCK with RAD51 and BRCA1 in a kinase dependent manner. Attenuation of LCK sensitized HR-proficient eEOC cells to PARP inhibitor in cell culture and pre-clinical mouse studies. These findings identify a new mechanism for expanding utility of PARP inhibitors in HR proficient ovarian cancer. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=120 SRC="FIGDIR/small/433791v2_ufig1.gif" ALT="Figure 1"> View larger version (33K): org.highwire.dtl.DTLVardef@1715c60org.highwire.dtl.DTLVardef@167cb5forg.highwire.dtl.DTLVardef@158d807org.highwire.dtl.DTLVardef@1ced201_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2021

Preprints in motion: tracking changes between posting and journal publication

Amidst the COVID-19 pandemic, preprints in the biomedical sciences are being posted and accessed at unprecedented rates, drawing widespread attention from the general public, press and policymakers for the first time. This phenomenon has sharpened longstanding questions about the reliability of information shared prior to journal peer review. Does the information shared in preprints typically withstand the scrutiny of peer review, or are conclusions likely to change in the version of record? We assessed preprints from bioRxiv and medRxiv that had been posted and subsequently published in a journal through 30th April 2020, representing the initial phase of the pandemic response. We utilised a combination of automatic and manual annotations to quantify how an article changed between the preprinted and published version. We found that the total number of figure panels and tables changed little between preprint and published articles. Moreover, the conclusions of 7.2% of non-COVID-19-related and 17.2% of COVID-19-related abstracts undergo a discrete change by the time of publication, but the majority of these changes do not qualitatively change the conclusions of the paper.

scientific communication and education↗

bioRxiv2020

Pretreatment with LCK Inhibitors Chemosensitizes Cisplatin-Resistant Endometrioid Ovarian Tumors

ObjectiveTo evaluate LCK inhibitors (LCKi) as chemosensitizing agents for platinum-resistant endometrioid ovarian carcinoma. MethodsKM Plotter survival data was obtained for endometrioid ovarian cancer based on LCK mRNA expression. Cisplatin resistant endometrioid ovarian carcinoma cell lines were cultured and treated first with LCKi or vehicle, then combination LCKi-cisplatin. Cell viability was assessed via CellTiter-Glo, and apoptosis with Caspase 3/7 Assay. Protein lysates were isolated from treated cells, with{gamma} -H2Ax, a DNA adduct marker, assessed. In vivo study compared mice treated with vehicle or LCK inhibitor followed by LCK inhibitor, cisplatin, or combination therapy. One-way ANOVA and two sample t-test were used to assess statistical significance with GraphPad Prism. ResultsKM plotter data indicated LCK expression is associated with significantly worse median progression-free survival (HR 3.19, p=0.02), and a trend toward decreased overall survival in endometrioid ovarian tumors with elevated LCK expression (HR 2.45, p=0.41). In vitro, cisplatin resistant ovarian endometrioid cells treated first with LCKi followed by combination LCKi-cisplatin treatment showed decreased cell viability and increased apoptosis. Immunoblot studies revealed inhibition of LCK led to increased expression of{gamma} -H2AX. In vivo results demonstrate treatment with LCKi followed by LCKi-cisplatin leads to significantly slowed tumor growth. ConclusionsWe identified a targetable pathway for chemosensitization of platinum resistant endometrioid ovarian cancer with initial treatment of LCKi followed by co-treatment with LCKi-cisplatin.

cancer biology↗

bioRxiv2020

Preprinting a pandemic: the role of preprints in the COVID-19 pandemic

The world continues to face a life-threatening viral pandemic. The virus underlying the COVID-19 disease, SARS-CoV-2, has caused over 98 million confirmed cases and 2.2 million deaths since January 2020. Although the most recent respiratory viral pandemic swept the globe only a decade ago, the way science operates and responds to current events has experienced a paradigm shift in the interim. The scientific community has responded rapidly to the COVID-19 pandemic, releasing over 125,000 COVID-19 related scientific articles within 10 months of the first confirmed case, of which more than 30,000 were hosted by preprint servers. We focused our analysis on bioRxiv and medRxiv, two growing preprint servers for biomedical research, investigating the attributes of COVID-19 preprints, their access and usage rates, as well as characteristics of their propagation on online platforms. Our data provides evidence for increased scientific and public engagement with preprints related to COVID-19 (COVID-19 preprints are accessed more, cited more, and shared more on various online platforms than non-COVID-19 preprints), as well as changes in the use of preprints by journalists and policymakers. We also find evidence for changes in preprinting and publishing behaviour: COVID-19 preprints are shorter and reviewed faster. Our results highlight the unprecedented role of preprints and preprint servers in the dissemination of COVID-19 science, and the impact of the pandemic on the scientific communication landscape.

scientific communication and education↗

bioRxiv2020

Live cell imaging of the hyperthermophilic archaeon Sulfolobus acidocaldarius identifies complementary roles for two ESCRTIII homologues in ensuring a robust and symmetric cell division

Live-cell imaging has revolutionized our understanding of dynamic cellular processes in bacteria and eukaryotes. While similar techniques have recently been applied to the study of halophilic archaea, our ability to explore the cell biology of thermophilic archaea is limited, due to the technical challenges of imaging at high temperatures. Here, we report the construction of the Sulfoscope, a heated chamber that enables live-cell imaging on an inverted fluorescent microscope. Using this system combined with thermostable fluorescent probes, we were able to image Sulfolobus cells as they divide, revealing a tight coupling between changes in DNA compaction, segregation and cytokinesis. By imaging deletion mutants, we observe important differences in the function of the two ESCRTIII proteins recently implicated in cytokinesis. The loss of CdvB1 compromises cell division, causing occasional division failures and fusion of the two daughter cells, whereas the deletion of cdvB2 leads to a profound loss of division symmetry, generating daughter cells that vary widely in size and eventually generating ghost cells. These data indicate that DNA separation and cytokinesis are coordinated in Sulfolobus, as is the case in eukaryotes, and that two contractile ESCRTIII polymers perform distinct roles to ensure that Sulfolobus cells undergo a robust and symmetrical division. Taken together, the Sulfoscope has shown to provide a controlled high temperature environment, in which cell biology of Sulfolobus can be studied in unprecedent details.

microbiology↗

bioRxiv2019

Closed mitosis requires local disassembly of the nuclear envelope

At the end of mitosis, eukaryotic cells must segregate both copies of their replicated genome into two new nuclear compartments (1). They do this either by first dismantling and later reassembling the nuclear envelope in a so called \"open mitosis\", or by reshaping an intact nucleus and then dividing into two in a \"closed mitosis\" (2, 3). However, while mitosis has been studied in a wide variety of eukaryotes for over a century (4), it is not known how the double membrane of the nuclear envelope is split into two at the end of a closed mitosis without compromising the impermeability of the nuclear compartment (5). In studying this problem in the fission yeast Schizosaccharomyces pombe, a classical model for closed mitosis (5), we use genetics, live cell imaging and electron tomography to show that nuclear fission is achieved via local disassembly of the nuclear envelope (NE) within the narrow bridge that links segregating daughter nuclei. In doing so, we identify a novel inner NE-localised protein Les1 that restricts the process of local NE breakdown (local NEB) to the bridge midzone and prevents the leakage of material from daughter nuclei. The mechanics of local NEB in a closed mitosis closely mirror those of NEB in open mitosis (3), revealing an unexpectedly deep conservation of nuclear remodelling mechanisms across diverse eukaryotes.

cell biology↗

bioRxiv2019

Proteasome-mediated protein degradation resets the cell division cycle and triggers ESCRT-III-mediated cytokinesis in an archaeon

The archaeon Sulfolobus acidocaldarius is a relative of eukaryotes known to progress orderly through its cell division cycle despite lacking obvious CDK/cyclin homologues. Here, in exploring the mechanisms underpinning archaeal cell division cycle control, we show that the proteasome of S. acidocaldarius, like its eukaryotic counterpart, regulates the transition from the end of one cell division cycle to the beginning of the next. Further, we identify the archaeal ESCRT-III homologue CdvB as a key target of the proteasome, and show that state-dependent degradation of CdvB triggers archaeal cell division by allowing constriction of a CdvB1:CdvB2 ESCRT-III division ring. These findings suggest an ancient role for proteasome-mediated degradation in resetting the cell division cycle in both archaea and eukaryotes.

cell biology↗

bioRxiv2019

Proteome-wide characterization and biomarker identification of intracranial aneurysm

The scientific basis of intracranial aneurysm (IA) formation, its rupture and further development of cerebral vasospasm remains incompletely understood. Deciphering the molecular mechanisms underlying these events will lead to identification of early detection biomarkers and in turn, improved treatment outcomes. Aberrant protein expression may drive structural alterations of vasculature found in IA. To unravel these aberrations, we performed untargeted, global, quantitative proteomic analysis of aneurysm tissue and serum from patients with IA. Samples were derived from patients of three clinical sub groups- 1) unruptured aneurysm 2) ruptured aneurysm without vasospasm 3) ruptured aneurysm who developed vasospasm. A total of 937 and 294 proteins were identified in aneurysm tissue and serum samples respectively. Several proteins that are known to maintain the structural integrity of vasculature were found to be dysregulated. ORM1, a glycoprotein, was significantly upregulated in both the aneurysm tissue and serum samples of unruptured IA patients. We employed a larger cohort of patients and validated ORM1 as a potential biomarker for screening of unruptured aneurysm using ELISA. Samples from ruptured aneurysm with vasospasm showed significant upregulation of MMP9 as compared to ruptured aneurysm without vasospasm. Using a cohort of ruptured aneurysm patients with and without vasospasm, we validated MMP9 as a potential biomarker for vasospasm. This study reveals pathophysiology underlying different clinical sub groups of IA and also suggests potential biomarkers.

neuroscience↗