40 research outputs found

    Mechanism of nuclear movements in a multinucleated cell.

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    Multinucleated cells are important in many organisms, but the mechanisms governing the movements of nuclei sharing a common cytoplasm are not understood. In the hyphae of the plant pathogenic fungus Ashbya gossypii, nuclei move back and forth, occasionally bypassing each other, preventing the formation of nuclear clusters. This is essential for genetic stability. These movements depend on cytoplasmic microtubules emanating from the nuclei that are pulled by dynein motors anchored at the cortex. Using three-dimensional stochastic simulations with parameters constrained by the literature, we predict the cortical anchor density from the characteristics of nuclear movements. The model accounts for the complex nuclear movements seen in vivo, using a minimal set of experimentally determined ingredients. Of interest, these ingredients power the oscillations of the anaphase spindle in budding yeast, but in A. gossypii, this system is not restricted to a specific nuclear cycle stage, possibly as a result of adaptation to hyphal growth and multinuclearity

    Geometrical and mechanical properties control actin filament organization.

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    The different actin structures governing eukaryotic cell shape and movement are not only determined by the properties of the actin filaments and associated proteins, but also by geometrical constraints. We recently demonstrated that limiting nucleation to specific regions was sufficient to obtain actin networks with different organization. To further investigate how spatially constrained actin nucleation determines the emergent actin organization, we performed detailed simulations of the actin filament system using Cytosim. We first calibrated the steric interaction between filaments, by matching, in simulations and experiments, the bundled actin organization observed with a rectangular bar of nucleating factor. We then studied the overall organization of actin filaments generated by more complex pattern geometries used experimentally. We found that the fraction of parallel versus antiparallel bundles is determined by the mechanical properties of actin filament or bundles and the efficiency of nucleation. Thus nucleation geometry, actin filaments local interactions, bundle rigidity, and nucleation efficiency are the key parameters controlling the emergent actin architecture. We finally simulated more complex nucleation patterns and performed the corresponding experiments to confirm the predictive capabilities of the model

    A quantitative map of nuclear pore assembly reveals two distinct mechanisms

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    Understanding how the nuclear pore complex (NPC) assembles is of fundamental importance to grasp the mechanisms behind its essential function and understand its role during evolution of eukaryotes1–4. While we know that at least two NPC assembly pathways exist, one during exit from mitosis and one during nuclear growth in interphase, we currently lack a quantitative map of their molecular events. Here, we use fluorescence correlation spectroscopy (FCS) calibrated live imaging of endogenously fluorescently-tagged nucleoporins to map the changes in composition and stoichiometry of seven major modules of the human NPC during its assembly in single dividing cells. This systematic quantitative map reveals that the two assembly pathways employ strikingly different molecular mechanisms, inverting the order of addition of two large structural components, the central ring complex and nuclear filaments. Our dynamic stoichiometry data allows us to perform the first computational simulation that predicts the structure of postmitotic NPC assembly intermediates

    Mammalian oocytes store proteins for the early embryo on cytoplasmic lattices

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    Mammalian oocytes are filled with poorly understood structures called cytoplasmic lattices. First discovered in the 1960s and speculated to correspond to mammalian yolk, ribosomal arrays, or intermediate filaments, their function has remained enigmatic to date. Here, we show that cytoplasmic lattices are sites where oocytes store essential proteins for early embryonic development. Using super-resolution light microscopy and cryoelectron tomography, we show that cytoplasmic lattices are composed of filaments with a high surface area, which contain PADI6 and subcortical maternal complex proteins. The lattices associate with many proteins critical for embryonic development, including proteins that control epigenetic reprogramming of the preimplantation embryo. Loss of cytoplasmic lattices by knocking out PADI6 or the subcortical maternal complex prevents the accumulation of these proteins and results in early embryonic arrest. Our work suggests that cytoplasmic lattices enrich maternally provided proteins to prevent their premature degradation and cellular activity, thereby enabling early mammalian development

    Nuclear pore assembles via structurally and molecularly distinct mechanisms after mitosis and during interphase

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    The nuclear pore complex (NPC) is the largest non-polymeric protein complex in eukaryotic cells and spans the double membrane of the nucleus (nuclear envelope; NE) to mediate nucleocytoplasmic transport. In mammalian cells, NPCs are assembled in two cell cycle stages, during nuclear assembly after mitosis and nuclear growth in interphase. How the NPC and the double nuclear membrane reassemble concomitantly in late mitosis, and how the NPC newly assembles in the closed NE in interphase, has been unclear. By correlating live imaging with three-dimensional electron microscopy, we have recently revealed that nuclear pores assemble via structurally distinct mechanisms in mitosis and interphase; during mitotic exit, pore assembly proceeds by radial dilation of small membrane openings, while in interphase, assembly induces a novel asymmetric inside-out fusion of the inner with the outer nuclear membrane. To understand the molecular maturation processes of these two distinct NPC assembly pathways, we created genome-edited GFP knock-in cells for nucleoporins of all major NPC substructures, i.e. the cytoplasmic filaments, the cytoplasmic/nucleoplasmic rings, the inner rings, and the nuclear basket. By FCS-calibrated three-dimensional live cell imaging, we monitored the concentration changes of these GFP-tagged nucleoporins in different regions of the NE where postmitotic and interphase assembly can be spatially distinguished for the first hour after mitotic exit. Quantitative kinetic analysis of the concentration changes showed that the molecular assembly order and maturation kinetics are distinct for postmitotic and interphase assembly, demonstrating that NPC assembly is not only a structurally but also molecularly different process between mitosis and interphas

    Collective light-matter interaction in the presence of atomic recoil

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    International audienceno abstrac

    Feedback, Mass Conservation and Reaction Kinetics Impact the Robustness of Cellular Oscillations

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    <div><p>Oscillations occur in a wide variety of cellular processes, for example in calcium and p53 signaling responses, in metabolic pathways or within gene-regulatory networks, e.g. the circadian system. Since it is of central importance to understand the influence of perturbations on the dynamics of these systems a number of experimental and theoretical studies have examined their robustness. The period of circadian oscillations has been found to be very robust and to provide reliable timing. For intracellular calcium oscillations the period has been shown to be very sensitive and to allow for frequency-encoded signaling. We here apply a comprehensive computational approach to study the robustness of period and amplitude of oscillatory systems. We employ different prototype oscillator models and a large number of parameter sets obtained by random sampling. This framework is used to examine the effect of three design principles on the sensitivities towards perturbations of the kinetic parameters. We find that a prototype oscillator with negative feedback has lower period sensitivities than a prototype oscillator relying on positive feedback, but on average higher amplitude sensitivities. For both oscillator types, the use of Michaelis-Menten instead of mass action kinetics in all degradation and conversion reactions leads to an increase in period as well as amplitude sensitivities. We observe moderate changes in sensitivities if replacing mass conversion reactions by purely regulatory reactions. These insights are validated for a set of established models of various cellular rhythms. Overall, our work highlights the importance of reaction kinetics and feedback type for the variability of period and amplitude and therefore for the establishment of predictive models.</p></div
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