Cell-sized liposomes reveal how actin-myosin cortical tension drives shape change

Abstract

Animal cells actively generate contractile stress in the actin cortex, a thin actin network beneath the cell membrane, to facilitate shape changes during processes like cytokinesis and motility. On the microscopic scale, this stress is generated by myosin molecular motors, which bind to actin cytoskeletal filaments and use chemical energy to exert pulling forces. To decipher the physical basis for the regulation of cell shape changes, here, we use a cell-like system with a cortex anchored to the outside or inside of a liposome membrane. This system enables us to dissect the interplay between motor pulling forces, cortex–membrane anchoring, and network connectivity. We show that cortices on the outside of liposomes either spontaneously rupture and relax built-up mechanical stress by peeling away around the liposome or actively compress and crush the liposome. The decision between peeling and crushing depends on the cortical tension determined by the amount of motors and also on the connectivity of the cortex and its attachment to the membrane. Membrane anchoring strongly affects the morphology of cortex contraction inside liposomes: cortices contract inward when weakly attached, whereas they contract toward the membrane when strongly attached. We propose a physical model based on a balance of active tension and mechanical resistance to rupture. Our findings show how membrane attachment and network connectivity are able to regulate actin cortex remodeling and membrane-shape changes for cell polarization.

Shape control of lipid bilayer membranes by confined actin bundles

Abstract

In living cells, lipid membranes and biopolymers determine each other’s conformation in a delicate force balance. Cellular polymers such as actin filaments are strongly confined by the plasma membrane in cell protrusions such as lamellipodia and filopodia. Conversely, protrusion formation is facilitated by actin-driven membrane deformation and these protrusions are maintained by dense actin networks or bundles of actin filaments. Here we investigate the mechanical interplay between actin bundles and lipid bilayer membranes by reconstituting a minimal model system based on cell-sized liposomes with encapsulated actin filaments bundled by fascin. To address the competition between the deformability of the membrane and the enclosed actin bundles, we tune the bundle stiffness (through the fascin-to-actin molar ratio) and the membrane rigidity (through protein decoration). Using confocal microscopy and quantitative image analysis, we show that actin bundles deform the liposomes into a rich set of morphologies. For liposomes having a small membrane bending rigidity, the actin bundles tend to generate finger-like membrane protrusions that resemble cellular filopodia. Stiffer bundles formed at high crosslink density stay straight in the liposome body, whereas softer bundles formed at low crosslink density are bent and kinked. When the membrane has a large bending rigidity, membrane protrusions are suppressed. In this case, membrane enclosure forces the actin bundles to organize into cortical rings, to minimize the energy cost associated with filament bending. Our results highlight the importance of taking into account mechanical interactions between the actin cytoskeleton and the membrane to understand cell shape control.

Wu Zhi-Qian

Violla Bassim

Members

Johan d’Humières

JUNIOR GROUP LEADER POSITION

logo1logo2

 

 

 

 

Institute Curie (https://institut-curie.org) comprises a world-class multidisciplinary research center combining research in cell biology, genetics, epigenetics, immunology, soft matter physics, organic and medicinal chemistry and a hospital. Highly international, it includes over 3,300 researchers, physicians, clinicians, technicians and administrative staff working on three sites: Paris, Orsay and Saint-Cloud. Its core facilities include advanced imaging technologies, high throughput sequencing, bioinformatics, and proteomics (https://science.institut-curie.org/platforms/) as well as access to large clinical databases and sample collections through the hospital. The Institute is engaged in international partnerships, including the EU-Life alliance (https://eu-life.eu/) and the LifeTime initiative (https://lifetime-initiative.eu/).

Institute Curie is supporting the recruitment of a junior group leader position on its research campus in central Paris in the “Nuclear Dynamics” unit (http://umr3664.curie.fr/en), which is part of the Labex DEEP for “Development, Epigenesis, Epigenetics and LifeTime potential” (https://labex-deep.institut-curie.org/). The five research groups of the unit investigate the mechanisms underlying the stability and the plasticity of genetic and epigenetic information in normal or pathological contexts during development and throughout evolution. Using complementary and interdisciplinary approaches, they aim at developing an integrated view of the functional organization of the genome at different scales: from the molecule to the cell and up to the whole organism.

We invite applications from outstanding candidates wishing to address questions on the fundamental mechanisms of nuclear organization, genome biology and epigenetics, through computational and/or modeling approaches. We are looking for candidates with a strong expertise in bioinformatics, computational biology and/or theoretical biological physics as well as a proven ability to collaborate with experimentalists. If needed, occasional access to wet lab space can be provided. Selection of the candidates will be based on the outstanding quality of their scientific record and project, their capacity to interface with other teams in the unit and beyond in the research center, and their strong collaborative spirit.

The newly recruited group will benefit from a start-up package from the Curie Research center and will be allocated a space adapted for 6 people. In addition, support from the Labex DEEP will complement the package. Successful candidates should meet criteria to compete for national and international funding, and for French institutional research positions (CNRS or University).


Send full CV, motivation letter, 3-4 pages research plan and contact details of 3-5 individuals who can be contacted for recommendation letters. Short listed candidates will be invited to present their past achievements and future research program.
Applications should be sent to: call.umr3664@curie.fr
For information please contact: Angela Taddei: angela.taddei@curie.fr, +33 156246704

Deadline for applications: March 15, 2021. Interviews scheduled in May 2021


Institut Curie is an inclusive, equal opportunities employer and is dedicated to the highest standards of research integrity.

logoHR

 

Competition of ligands and the 18-mer binding domain of the RHAU helicase for G-quadruplexes

Stabilizing particular DNA and RNA structures called G-quadruplexes (G4s) using specific ligands (L) is a strategy proposed to fight cancer. However, while G4:L interactions are often investigated, whether or not ligands are able to disrupt interactions between G4s and proteins (P) remains poorly studied. Here, using native mass spectrometry, we investigated ternary G4:L:P complexes formed by G4s, some of the highest affinity ligands, and the binding domain of the RHAU helicase. First, our results suggest that RHAU binds not only preferentially to parallel G4s but to free external G-quartets. We also found that, depending on the G4, ligands could prevent the binding of the peptide, either by direct competition for the binding sites (orthosteric inhibition) or by inducing conformational changes (allosteric inhibition). Notably, the ligand Cu-ttpy induced a conformational change that increased the binding of the peptide. This study illustrates that it is important to not only characterize drug-target interactions but also how the binding to other partners is affected.

Three-component C-H bond sulfonylation of imidazoheterocycles via visible-light organophotoredox

The first entirely visible-light photoredox catalyzed sulfonylation of imidazoheterocycles has been developed. This transformation demonstrates an efficient C-H functionalization for the straightforward synthesis of novel C-3 sulfonylated imidazoheterocycles from various imidazopyridines and diaryliodonium salts with different electronic and steric properties and easy handled DABCO- bis (sulfur dioxide). The reaction proceeds in moderate to good yields under mild conditions at room temperature using the inexpensive organophotocatalyst EosinY.Na 2 and shows a high functional group tolerance (37 examples).

Drice Challal

Amoeboid Swimming Is Propelled by Molecular Paddling in Lymphocytes.

Mammalian cells developed two main migration modes. The slow mesenchymatous mode, like crawling of fibroblasts, relies on maturation of adhesion complexes and actin fiber traction, whereas the fast amoeboid mode, observed exclusively for leukocytes and cancer cells, is characterized by weak adhesion, highly dynamic cell shapes, and ubiquitous motility on two-dimensional and in three-dimensional solid matrix. In both cases, interactions with the substrate by adhesion or friction are widely accepted as a prerequisite for mammalian cell motility, which precludes swimming. We show here experimental and computational evidence that leukocytes do swim, and that efficient propulsion is not fueled by waves of cell deformation but by a rearward and inhomogeneous treadmilling of the cell external membrane. Our model consists of a molecular paddling by transmembrane proteins linked to and advected by the actin cortex, whereas freely diffusing transmembrane proteins hinder swimming. Furthermore, continuous paddling is enabled by a combination of external treadmilling and selective recycling by internal vesicular transport of cortex-bound transmembrane proteins. This mechanism explains observations that swimming is five times slower than the retrograde flow of cortex and also that lymphocytes are motile in nonadherent confined environments. Resultantly, the ubiquitous ability of mammalian amoeboid cells to migrate in two dimensions or three dimensions and with or without adhesion can be explained for lymphocytes by a single machinery of heterogeneous membrane treadmilling.

Extracellular vesicles and chronic inflammation during HIV infection.

Inflammation is a hallmark of HIV infection. Among the multiple stimuli that can induce inflammation in untreated infection, ongoing viral replication is a primary driver. After initiation of effective combined antiretroviral therapy (cART), HIV replication is drastically reduced or halted. However, even virologically controlled patients may continue to have abnormal levels of inflammation. A number of factors have been proposed to cause inflammation in HIV infection: among others, residual (low-level) HIV replication, production of HIV protein or RNA in the absence of replication, microbial translocation from the gut to the circulation, co-infections, and loss of immunoregulatory responses. Importantly, chronic inflammation in HIV-infected individuals increases the risk for a number of non-infectious co-morbidities, including cancer and cardiovascular disease. Thus, achieving a better understanding of the underlying mechanisms of HIV-associated inflammation in the presence of cART is of utmost importance. Extracellular vesicles have emerged as novel actors in intercellular communication, involved in a myriad of physiological and pathological processes, including inflammation. In this review, we will discuss the role of extracellular vesicles in the pathogenesis of HIV infection, with particular emphasis on their role as inducers of chronic inflammation.

Extracellular vesicles or exosomes? On primacy, precision, and popularity influencing a choice of

Journal of extracellular vesicles: the seven year itch!

SnapShot: Extracellular Vesicles.

Cells release a variety of extracellular vesicles (EVs; including exosomes, microvesicles, and many others) into their environment. EVs can bud in endosomes or directly at the plasma membrane, carrying a selection of components from the cell and displaying various functional properties. Different techniques can be used to separate EV subtypes and EVs from co-isolated components, resulting in preparations of different abundance and purity.

Therapy statement on extracellular vesicles from mesenchymal stromal cells and other cells: considerations

The International Society for Cellular and Gene Therapies (ISCT) and the International Society for Extracellular Vesicles (ISEV) recognize the potential of extracellular vesicles (EVs, including exosomes) from mesenchymal stromal cells (MSCs) and possibly other cell sources as treatments for COVID-19. Research and trials in this area are encouraged. However, ISEV and ISCT do not currently endorse the use of EVs or exosomes for any purpose in COVID-19, including but not limited to reducing cytokine storm, exerting regenerative effects or delivering drugs, pending the generation of appropriate manufacturing and quality control provisions, pre-clinical safety and efficacy data, rational clinical trial design and proper regulatory oversight.

Extracellular vesicles: eat glutamine and spit acidic bubbles.

Extracellular vesicles mediate transfer of diverse molecular content to target cells in order to induce phenotypic changes, which has put them under the spotlight as likely major players in cell-to-cell communication. However, extracellular vesicle heterogeneity in terms of intracellular origin has only recently been recognized as a potential determinant of their activity. Recent work by Fan et al (2020) illustrates how lack of external resources that affect cellular homeostasis and signaling can also modulate EV biogenesis, by inducing the production of a novel subpopulation of exosomes enriched in Rab11a with context-dependent roles in Drosophila gland physiology and cancer cell aggressiveness.

Methods for Separation and Characterization of Extracellular Vesicles: Results of a Worldwide

Research on extracellular vesicles (EVs) is growing exponentially due to an increasing appreciation of EVs as disease biomarkers and therapeutics, an expanding number of EV-containing materials under study, and application of new preparation, detection, and cargo analysis methods. Diversity of both sources and methodologies imposes challenges on the comparison of measurement results between studies and laboratories. While reference guidelines and minimal requirements for EV research have achieved the important objective of assembling community consensus, it is also essential to understand which methodologies and quality controls are currently being applied, and how usage trends are evolving. As an initial response to this need, the International Society for Extracellular Vesicles (ISEV) performed a worldwide survey in 2015 on « Techniques used for the isolation and characterization of extracellular vesicles » and published the results from this survey in 2016. In 2019, a new survey was performed to assess the changing state of the field. The questionnaire received more than 600 full or partial responses, and the present manuscript summarizes the results of this second worldwide survey. The results emphasize that separation methods such as ultracentrifugation and density gradients are still the most commonly used methods, the use of size exclusion chromatography has increased, and techniques based on tangential flow and microfluidics are now being used by more than 10% of respondents. The survey also reveals that most EV researchers still do not perform sample quality controls before or after isolation of EVs. Finally, the majority of EV researchers emphasize that separation and characterization of EVs should receive more attention.

CD163 tumor-associated macrophage accumulation in breast cancer patients reflects both local

The accumulation of tumor-associated macrophages (TAMs) is correlated with poor clinical outcome, but the mechanisms governing their differentiation from circulating monocytes remain unclear in humans.

Effects of interleukin-2 in immunostimulation and immunosuppression.

Historically, interleukin-2 (IL-2) was first described as an immunostimulatory factor that supports the expansion of activated effector T cells. A layer of sophistication arose when regulatory CD4+ T lymphocytes (Tregs) were shown to require IL-2 for their development, homeostasis, and immunosuppressive functions. Fundamental distinctions in the nature and spatiotemporal expression patterns of IL-2 receptor subunits on naive/memory/effector T cells versus Tregs are now being exploited to manipulate the immunomodulatory effects of IL-2 for therapeutic purposes. Although high-dose IL-2 administration has yielded discrete clinical responses, low-dose IL-2 as well as innovative strategies based on IL-2 derivatives, including « muteins, » immunocomplexes, and immunocytokines, are being explored to therapeutically enhance or inhibit the immune response.