The bacterial division apparatus catalyses the synthesis and remodelling of septal peptidoglycan (sPG) to build the cell wall layer that fortifies the daughter cell poles. Understanding of this essential process has been limited by the lack of native three-dimensional views of developing septa. Here, we apply state-of-the-art cryogenic electron tomography (cryo-ET) and fluorescence microscopy to visualize the division site architecture and sPG biogenesis dynamics of the Gram-negative bacterium Escherichia coli. We identify a wedge-like sPG structure that fortifies the ingrowing septum. Experiments with strains defective in sPG biogenesis revealed that the septal architecture and mode of division can be modified to more closely resemble that of other Gram-negative (Caulobacter crescentus) or Gram-positive (Staphylococcus aureus) bacteria, suggesting that a conserved mechanism underlies the formation of different septal morphologies. Finally, analysis of mutants impaired in amidase activation (ΔenvC ΔnlpD) showed that cell wall remodelling affects the placement and stability of the cytokinetic ring. Taken together, our results support a model in which competition between the cell elongation and division machineries determines the shape of cell constrictions and the poles they form. They also highlight how the activity of the division system can be modulated to help generate the diverse array of shapes observed in the bacterial domain.

• MeSH
• amidohydrolasy MeSH
• buněčná stěna MeSH
• buněčné dělení MeSH
• Escherichia coli * fyziologie   MeSH
• peptidoglykan * MeSH
• tvar buňky MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH

Plant cell morphogenesis involves concerted rearrangements of microtubules and actin microfilaments. We previously reported that FH1, the main Arabidopsis thaliana housekeeping Class I membrane-anchored formin, contributes to actin dynamics and microtubule stability in rhizodermis cells. Here we examine the effects of mutations affecting FH1 (At3g25500) on cell morphogenesis and above-ground organ development in seedlings, as well as on cytoskeletal organization and dynamics, using a combination of confocal and variable angle epifluorescence microscopy with a pharmacological approach. Homozygous fh1 mutants exhibited cotyledon epinasty and had larger cotyledon pavement cells with more pronounced lobes than the wild type. The pavement cell shape alterations were enhanced by expression of the fluorescent microtubule marker GFP-microtubule-associated protein 4 (MAP4). Mutant cotyledon pavement cells exhibited reduced density and increased stability of microfilament bundles, as well as enhanced dynamics of microtubules. Analogous results were also obtained upon treatments with the formin inhibitor SMIFH2 (small molecule inhibitor of formin homology 2 domains). Pavement cell shape in wild-type (wt) and fh1 plants in some situations exhibited a differential response towards anti-cytoskeletal drugs, especially the microtubule disruptor oryzalin. Our observations indicate that FH1 participates in the control of microtubule dynamics, possibly via its effects on actin, subsequently influencing cell morphogenesis and macroscopic organ development.

• MeSH
• aktiny metabolismus   MeSH
• Arabidopsis cytologie   účinky léků   metabolismus   MeSH
• biologické markery metabolismus   MeSH
• biologické modely MeSH
• cytoskelet účinky léků   metabolismus   MeSH
• fluorescence MeSH
• klathrin metabolismus   MeSH
• kotyledon účinky léků   metabolismus   MeSH
• membránové proteiny metabolismus   MeSH
• mikrofilamenta účinky léků   metabolismus   MeSH
• mikrotubuly účinky léků   metabolismus   MeSH
• mutace genetika   MeSH
• proteiny huseníčku metabolismus   MeSH
• semenáček účinky léků   růst a vývoj   metabolismus   MeSH
• thioketony farmakologie   MeSH
• tvar buňky * účinky léků   MeSH
• uracil analogy a deriváty   farmakologie   MeSH
• zelené fluorescenční proteiny metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

We present a fast and robust approach to tracking the evolving shape of whole fluorescent cells in time-lapse series. The proposed tracking scheme involves two steps. First, coherence-enhancing diffusion filtering is applied on each frame to reduce the amount of noise and enhance flow-like structures. Second, the cell boundaries are detected by minimizing the Chan-Vese model in the fast level set-like and graph cut frameworks. To allow simultaneous tracking of multiple cells over time, both frameworks have been integrated with a topological prior exploiting the object indication function. The potential of the proposed tracking scheme and the advantages and disadvantages of both frameworks are demonstrated on 2-D and 3-D time-lapse series of rat adipose-derived mesenchymal stem cells and human lung squamous cell carcinoma cells, respectively.

• MeSH
• buněčné jádro chemie   MeSH
• buněčný tracking metody   MeSH
• fluorescenční mikroskopie metody   MeSH
• krysa rodu rattus MeSH
• lidé MeSH
• mezenchymální kmenové buňky cytologie   MeSH
• nádorové buněčné linie MeSH
• počítačové zpracování obrazu metody   MeSH
• tvar buňky fyziologie   MeSH
• zvířata MeSH
• Check Tag
• krysa rodu rattus MeSH
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Nanoparticle-cell interactions begin with the cellular uptake of the nanoparticles, a process that eventually determines their cellular fate. In the present work, we show that the morphological features of nanodiamonds (NDs) affect both the anchoring and internalization stages of their endocytosis. While a prickly ND (with sharp edges/corners) has no trouble of anchoring onto the plasma membrane, it suffers from difficult internalization afterwards. In comparison, the internalization of a round ND (obtained by selective etching of the prickly ND) is not limited by its lower anchoring amount and presents a much higher endocytosis amount. Molecular dynamics simulation and continuum modelling results suggest that the observed difference in the anchoring of round and prickly NDs likely results from the reduced contact surface area with the cell membrane of the former, while the energy penalty associated with membrane curvature generation, which is lower for a round ND, may explain its higher probability of the subsequent internalization.

• MeSH
• biologické modely MeSH
• buněčná membrána chemie   MeSH
• buňky Hep G2 MeSH
• endocytóza MeSH
• HeLa buňky MeSH
• lidé MeSH
• nanodiamanty chemie   MeSH
• simulace molekulární dynamiky MeSH
• viabilita buněk MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

• MeSH
• entomologie MeSH
• fyziologie buňky MeSH
• hematologie MeSH
• kompartmentace buňky MeSH
• nádorové procesy MeSH
• Publikační typ
• kongresy MeSH

• Konspekt
• Biologické vědy
• NLK Obory
• biologie
• biologie
• onkologie

Overtly self-reactive T cells are removed during thymic selection. However, it has been recently established that T cell self-reactivity promotes protective immune responses. Apparently, the level of self-reactivity of mature T cells must be tightly balanced. Our mathematical model and experimental data show that the dynamic regulation of CD4- and CD8-LCK coupling establish the self-reactivity of the peripheral T cell pool. The stoichiometry of the interaction between CD8 and LCK, but not between CD4 and LCK, substantially increases upon T cell maturation. As a result, peripheral CD8+ T cells are more self-reactive than CD4+ T cells. The different levels of self-reactivity of mature CD8+ and CD4+ T cells likely reflect the unique roles of these subsets in immunity. These results indicate that the evolutionary selection pressure tuned the CD4-LCK and CD8-LCK stoichiometries, as they represent the unique parts of the proximal T cell receptor (TCR) signaling pathway, which differ between CD4+ and CD8+ T cells.

• MeSH
• antigeny metabolismus   MeSH
• buněčná diferenciace MeSH
• CD4-pozitivní T-lymfocyty cytologie   metabolismus   MeSH
• CD8-pozitivní T-lymfocyty cytologie   metabolismus   MeSH
• homeostáza MeSH
• myši inbrední C57BL MeSH
• signální transdukce MeSH
• tyrosinkinasa p56(lck), specifická pro lymfocyty metabolismus   MeSH
• vazba proteinů MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Intracellular and extracellular mechanical forces play a crucial role during tissue growth, modulating nuclear shape and function and resulting in complex collective cell behaviour. However, the mechanistic understanding of how the orientation, shape, symmetry and homogeneity of cells are affected by environmental geometry is still lacking. Here we investigate cooperative cell behaviour and patterns under geometric constraints created by topographically patterned substrates. We show how cells cooperatively adopt their geometry, shape, positioning of the nucleus and subsequent proliferation activity. Our findings indicate that geometric constraints induce significant squeezing of cells and nuclei, cytoskeleton reorganization, drastic condensation of chromatin resulting in a change in the cell proliferation rate and the anisotropic growth of cultures. Altogether, this work not only demonstrates complex non-trivial collective cellular responses to geometrical constraints but also provides a tentative explanation of the observed cell culture patterns grown on different topographically patterned substrates. These findings provide important fundamental knowledge, which could serve as a basis for better controlled tissue growth and cell-engineering applications.

• MeSH
• biologické modely * MeSH
• buněčné jádro fyziologie   ultrastruktura   MeSH
• buněčný převod mechanických signálů fyziologie   MeSH
• buňky Hep G2 MeSH
• lidé MeSH
• mezibuněčná komunikace fyziologie   MeSH
• počítačová simulace MeSH
• proliferace buněk fyziologie   MeSH
• velikost buňky * MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Data-driven cell tracking and segmentation methods in biomedical imaging require diverse and information-rich training data. In cases where the number of training samples is limited, synthetic computer-generated data sets can be used to improve these methods. This requires the synthesis of cell shapes as well as corresponding microscopy images using generative models. To synthesize realistic living cell shapes, the shape representation used by the generative model should be able to accurately represent fine details and changes in topology, which are common in cells. These requirements are not met by 3D voxel masks, which are restricted in resolution, and polygon meshes, which do not easily model processes like cell growth and mitosis. In this work, we propose to represent living cell shapes as level sets of signed distance functions (SDFs) which are estimated by neural networks. We optimize a fully-connected neural network to provide an implicit representation of the SDF value at any point in a 3D+time domain, conditioned on a learned latent code that is disentangled from the rotation of the cell shape. We demonstrate the effectiveness of this approach on cells that exhibit rapid deformations (Platynereis dumerilii), cells that grow and divide (C. elegans), and cells that have growing and branching filopodial protrusions (A549 human lung carcinoma cells). A quantitative evaluation using shape features and Dice similarity coefficients of real and synthetic cell shapes shows that our model can generate topologically plausible complex cell shapes in 3D+time with high similarity to real living cell shapes. Finally, we show how microscopy images of living cells that correspond to our generated cell shapes can be synthesized using an image-to-image model.

• MeSH
• Caenorhabditis elegans * MeSH
• lidé MeSH
• mitóza MeSH
• nádory plic * MeSH
• neuronové sítě MeSH
• počítačové zpracování obrazu metody   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

Cartilaginous structures are at the core of embryo growth and shaping before the bone forms. Here we report a novel principle of vertebrate cartilage growth that is based on introducing transversally-oriented clones into pre-existing cartilage. This mechanism of growth uncouples the lateral expansion of curved cartilaginous sheets from the control of cartilage thickness, a process which might be the evolutionary mechanism underlying adaptations of facial shape. In rod-shaped cartilage structures (Meckel, ribs and skeletal elements in developing limbs), the transverse integration of clonal columns determines the well-defined diameter and resulting rod-like morphology. We were able to alter cartilage shape by experimentally manipulating clonal geometries. Using in silico modeling, we discovered that anisotropic proliferation might explain cartilage bending and groove formation at the macro-scale.

• MeSH
• biologické modely MeSH
• chrupavka embryologie   MeSH
• myši MeSH
• obratlovci embryologie   MeSH
• počítačová simulace MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

BACKGROUND: Although the etiology of chronic lymphocytic leukemia (CLL), the most common type of adult leukemia, is still unclear, strong evidence implicates antigen involvement in disease ontogeny and evolution. Primary and 3D structure analysis has been utilised in order to discover indications of antigenic pressure. The latter has been mostly based on the 3D models of the clonotypic B cell receptor immunoglobulin (BcR IG) amino acid sequences. Therefore, their accuracy is directly dependent on the quality of the model construction algorithms and the specific methods used to compare the ensuing models. Thus far, reliable and robust methods that can group the IG 3D models based on their structural characteristics are missing. RESULTS: Here we propose a novel method for clustering a set of proteins based on their 3D structure focusing on 3D structures of BcR IG from a large series of patients with CLL. The method combines techniques from the areas of bioinformatics, 3D object recognition and machine learning. The clustering procedure is based on the extraction of 3D descriptors, encoding various properties of the local and global geometrical structure of the proteins. The descriptors are extracted from aligned pairs of proteins. A combination of individual 3D descriptors is also used as an additional method. The comparison of the automatically generated clusters to manual annotation by experts shows an increased accuracy when using the 3D descriptors compared to plain bioinformatics-based comparison. The accuracy is increased even more when using the combination of 3D descriptors. CONCLUSIONS: The experimental results verify that the use of 3D descriptors commonly used for 3D object recognition can be effectively applied to distinguishing structural differences of proteins. The proposed approach can be applied to provide hints for the existence of structural groups in a large set of unannotated BcR IG protein files in both CLL and, by logical extension, other contexts where it is relevant to characterize BcR IG structural similarity. The method does not present any limitations in application and can be extended to other types of proteins.

• MeSH
• anotace sekvence MeSH
• automatizace MeSH
• chronická lymfatická leukemie metabolismus   MeSH
• databáze proteinů MeSH
• imunoglobuliny chemie   MeSH
• lidé MeSH
• sekvence aminokyselin MeSH
• zobrazování trojrozměrné * MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH