Compression-dependent microtubule reinforcement enables cells to navigate confined environments.

Cells migrating through complex three-dimensional environments experience considerable physical challenges, including tensile stress and compression. To move, cells need to resist these forces while also squeezing the large nucleus through confined spaces. This requires highly coordinated cortical contractility. Microtubules can both resist compressive forces and sequester key actomyosin regulators to ensure appropriate activation of contractile forces. Yet, how these two roles are integrated to achieve nuclear transmigration in three dimensions is largely unknown. Here, we demonstrate that compression triggers reinforcement of a dedicated microtubule structure at the rear of the nucleus by the mechanoresponsive recruitment of cytoplasmic linker-associated proteins, which dynamically strengthens and repairs the lattice. These reinforced microtubules form the mechanostat: an adaptive feedback mechanism that allows the cell to both withstand compressive force and spatiotemporally organize contractility signalling pathways. The microtubule mechanostat facilitates nuclear positioning and coordinates force production to enable the cell to pass through constrictions. Disruption of the mechanostat imbalances cortical contractility, stalling migration and ultimately resulting in catastrophic cell rupture. Our findings reveal a role for microtubules as cellular sensors that detect and respond to compressive forces, enabling movement and ensuring survival in mechanically demanding environments.

Rigidity percolation and active advection synergize in the actomyosin cortex to drive amoeboid cell motility.

Spontaneous locomotion is a common feature of most metazoan cells, generally attributed to the properties of actomyosin networks. This force-producing machinery has been studied down to the most minute molecular details, especially in lamellipodium-driven migration. Nevertheless, how actomyosin networks work inside contraction-driven amoeboid cells still lacks unifying principles. Here, using stable motile blebs from HeLa cells as a model amoeboid motile system, we imaged the dynamics of the actin cortex at the single filament level and revealed the co-existence of three distinct rheological phases. We introduce "advected percolation," a process where rigidity percolation and active advection synergize, spatially organizing the actin network's mechanical properties into a minimal and generic locomotion mechanism. Expanding from our observations on simplified systems, we speculate that this model could explain, down to the single actin filament level, how amoeboid cells, such as cancer or immune cells, can propel efficiently through complex 3D environments.

Form follows function: Nuclear morphology as a quantifiable predictor of cellular senescence.

Enlarged or irregularly shaped nuclei are frequently observed in tissue cells undergoing senescence. However, it remained unclear whether this peculiar morphology is a cause or a consequence of senescence and how informative it is in distinguishing between proliferative and senescent cells. Recent research reveals that nuclear morphology can act as a predictive biomarker of senescence, suggesting an active role for the nucleus in driving senescence phenotypes. By employing deep learning algorithms to analyze nuclear morphology, accurate classification of cells as proliferative or senescent is achievable across various cell types and species both in vitro and in vivo. This quantitative imaging-based approach can be employed to establish links between senescence burden and clinical data, aiding in the understanding of age-related diseases, as well as assisting in disease prognosis and treatment response.

Nuclear fragility, blaming the blebs.

Although textbook pictures depict the cell nucleus as a simple ovoid object, it is now clear that it adopts a large variety of shapes in tissues. When cells deform, because of cell crowding or migration through dense matrices, the nucleus is subjected to large constraints that alter its shape. In this review, we discuss recent studies related to nuclear fragility, focusing on the surprising finding that the nuclear envelope can form blebs. Contrary to the better-known plasma membrane blebs, nuclear blebs are unstable and almost systematically lead to nuclear envelope opening and uncontrolled nucleocytoplasmic mixing. They expand, burst, and repair repeatedly when the nucleus is strongly deformed. Although blebs are a major source of nuclear instability, they are poorly understood so far, which calls for more in-depth studies of these structures.

RhoG deficiency abrogates cytotoxicity of human lymphocytes and causes hemophagocytic lymphohistiocytosis.

Exocytosis of cytotoxic granules (CG) by lymphocytes is required for the elimination of infected and malignant cells. Impairments in this process underly a group of diseases with dramatic hyperferritinemic inflammation termed hemophagocytic lymphohistiocytosis (HLH). Although genetic and functional studies of HLH have identified proteins controlling distinct steps of CG exocytosis, the molecular mechanisms that spatiotemporally coordinate CG release remain partially elusive. We studied a patient exhibiting characteristic clinical features of HLH associated with markedly impaired cytotoxic T lymphocyte (CTL) and natural killer (NK) cell exocytosis functions, who beared biallelic deleterious mutations in the gene encoding the small GTPase RhoG. Experimental ablation of RHOG in a model cell line and primary CTLs from healthy individuals uncovered a hitherto unappreciated role of RhoG in retaining CGs in the vicinity of the plasma membrane (PM), a fundamental prerequisite for CG exocytotic release. We discovered that RhoG engages in a protein-protein interaction with Munc13-4, an exocytosis protein essential for CG fusion with the PM. We show that this interaction is critical for docking of Munc13-4+ CGs to the PM and subsequent membrane fusion and release of CG content. Thus, our study illuminates RhoG as a novel essential regulator of human lymphocyte cytotoxicity and provides the molecular pathomechanism behind the identified here and previously unreported genetically determined form of HLH.

The cytoskeletal regulator HEM1 governs B cell development and prevents autoimmunity.

The WAVE regulatory complex (WRC) is crucial for assembly of the peripheral branched actin network constituting one of the main drivers of eukaryotic cell migration. Here, we uncover an essential role of the hematopoietic-specific WRC component HEM1 for immune cell development. Germline-encoded HEM1 deficiency underlies an inborn error of immunity with systemic autoimmunity, at cellular level marked by WRC destabilization, reduced filamentous actin, and failure to assemble lamellipodia. Hem1-/- mice display systemic autoimmunity, phenocopying the human disease. In the absence of Hem1, B cells become deprived of extracellular stimuli necessary to maintain the strength of B cell receptor signaling at a level permissive for survival of non-autoreactive B cells. This shifts the balance of B cell fate choices toward autoreactive B cells and thus autoimmunity.

Meeting report - Building the Cell 2018.

Cell biologists from all around the world gathered in Paris on the 26 to 28 September 2018 to participate in the 3rd international meeting 'Building the Cell'. It was organized by Hélène Barelli, Arnaud Echard, Thierry Galli, Florence Niedergang, Manuel Théry and Marie Hélène Verlhac on behalf of the French Society for Cell Biology (SBCF) at the Institut Pasteur. Around 230 participants joined the meeting for stimulating talks, discussions, poster sessions, and a gala dinner on the Seine that included a music performance by the rock group 'Membrane Band'. The unifying theme of the meeting was the development of creative multidisciplinary approaches to understand cellular life at different scales in a dynamic and quantitative manner. Here, we summarize the results presented at the meeting and the emerging ideas from the different sessions.

The "nuclear physics" behind epigenetic control of cell fate.

It is well understood that replicative and transcriptional responses in the nucleus occur under the influence of specific extracellular biochemical signals (e.g. growth factors and cytokines). However, it has become apparent recently that the nucleus is also able to sense and respond to more generic cues, such as physical forces and mechanical constraints. Indeed, being the largest and stiffest intracellular organelle, the nucleus is exposed to various types of forces acting from inside and outside the cell. These forces result in global and local deformations of the nucleus, which can significantly affect spatial organization and mechanical state of the nuclear envelope (NE). Considering that peripheral chromatin is attached to the NE, forces applied to the NE are transmitted to chromatin. This, in turn, can impact chromatin organization, dynamics, and activity. Where do these forces originate from and what are the physiological contexts in which they modulate critical nuclear activities? Discussing these questions is the main goal of the present mini-review.

Forcing Entry into the Nucleus.

Nuclear pore complexes tightly regulate nucleo-cytoplasmic transport, controlling the nuclear concentration of several transcription factors. In a recent issue of Cell, Elosegui-Artola et al. (2017) show that nuclear deformation modulates the nuclear entry rates of YAP/TAZ via nuclear pore stretching, clarifying how forces affect gene transcription.

Competition for actin between two distinct F-actin networks defines a bistable switch for cell polarization.

Symmetry-breaking polarization enables functional plasticity of cells and tissues and is yet not well understood. Here we show that epithelial cells, hard-wired to maintain a static morphology and to preserve tissue organization, can spontaneously switch to a migratory polarized phenotype after relaxation of the actomyosin cytoskeleton. We find that myosin II engages actin in the formation of cortical actomyosin bundles and thus makes it unavailable for deployment in the process of dendritic growth normally driving cell motility. Under low-contractility regimes, epithelial cells polarize in a front-back manner owing to the emergence of actin retrograde flows powered by dendritic polymerization of actin. Coupled to cell movement, the flows transport myosin II from the front to the back of the cell, where the motor locally 'locks' actin in contractile bundles. This polarization mechanism could be employed by embryonic and cancer epithelial cells in microenvironments where high-contractility-driven cell motion is inefficient.

Stimulation of the CLIP-170--dependent capture of membrane organelles by microtubules through fine tuning of microtubule assembly dynamics.

Cytoplasmic microtubules (MTs) continuously grow and shorten at their free plus ends, a behavior that allows them to capture membrane organelles destined for MT minus end-directed transport. In Xenopus melanophores, the capture of pigment granules (melanosomes) involves the +TIP CLIP-170, which is enriched at growing MT plus ends. Here we used Xenopus melanophores to test whether signals that stimulate minus end MT transport also enhance CLIP-170-dependent binding of melanosomes to MT tips. We found that these signals significantly (>twofold) increased the number of growing MT plus ends and their density at the cell periphery, thereby enhancing the likelihood of interaction with dispersed melanosomes. Computational simulations showed that local and global increases in the density of CLIP-170-decorated MT plus ends could reduce the half-time of melanosome aggregation by ~50%. We conclude that pigment granule aggregation signals in melanophores stimulate MT minus end-directed transport by the increasing number of growing MT plus ends decorated with CLIP-170 and redistributing these ends to more efficiently capture melanosomes throughout the cytoplasm.

Microtubules govern stress granule mobility and dynamics.

Stress granules (SGs) are ribonucleoprotein (RNP)-containing assemblies that are formed in the cytoplasm in response to stress. Previously, we demonstrated that microtubule depolymerization inhibited SG formation. Here, we show that arsenate-induced SGs move throughout the cytoplasm in a microtubule-dependent manner, and microtubules are required for SG disassembly, but not for SG persistence. Analysis of SG movement revealed that SGs exhibited obstructed diffusion on an average, though sometimes SGs demonstrated rapid displacements. Microtubule depolymerization did not influence preformed SG number and size, but significantly reduced the average velocity of SG movement, the frequency of quick movement events, and the apparent diffusion coefficient of SGs. Actin filament disruption had no effect on the SG motility. In cycloheximide-treated cells SGs dissociated into constituent parts that then dissolved within the cytoplasm. Microtubule depolymerization inhibited cycloheximide-induced SG disassembly. However, microtubule depolymerization did not influence the dynamics of poly(A)-binding protein (PABP) in SGs, according to FRAP results. We suggest that the increase of SG size is facilitated by the transport of smaller SGs along microtubules with subsequent fusion of them. At least some protein components of SGs can exchange with the cytoplasmic pool independently of microtubules.

CLIP-170-dependent capture of membrane organelles by microtubules initiates minus-end directed transport.

Cytoplasmic microtubules (MTs) continuously grow and shorten at free plus ends. During mitosis, this dynamic behavior allows MTs to capture chromosomes to initiate their movement to the spindle poles; however, the role of MT dynamics in capturing organelles for transport in interphase cells has not been demonstrated. Here we use Xenopus melanophores to test the hypothesis that MT dynamics significantly contribute to the efficiency of MT minus-end directed transport of membrane organelles. We demonstrate that initiation of transport of membrane-bounded melanosomes (pigment granules) to the cell center involves their capture by MT plus ends, and that inhibition of MT dynamics or loss of the MT plus-end tracking protein CLIP-170 from MT tips dramatically inhibits pigment aggregation. We conclude that MT dynamics are required for the initiation of MT transport of membrane organelles in interphase cells, and that +TIPs such as CLIP-170 play an important role in this process.