F-actin Assembly Research Articles

Loss of the actin regulator cyclase-associated protein 1 (CAP1) modestly affects dendritic spine remodeling during synaptic plasticity

Dendritic spines form the postsynaptic compartment of most excitatory synapses in the vertebrate brain. Morphological changes of dendritic spines contribute to major forms of synaptic plasticity such as long-term potentiation (LTP) or depression (LTD). Synaptic plasticity underlies learning and memory, and defects in synaptic plasticity contribute to the pathogeneses of human brain disorders. Hence, deciphering the molecules that drive spine remodeling during synaptic plasticity is critical for understanding the neuronal basis of physiological and pathological brain function. Since actin filaments (F-actin) define dendritic spine morphology, actin-binding proteins (ABP) that accelerate dis-/assembly of F-actin moved into the focus as critical regulators of synaptic plasticity. We recently identified cyclase-associated protein 1 (CAP1) as a novel actin regulator in neurons that cooperates with cofilin1, an ABP relevant for synaptic plasticity. We therefore hypothesized a crucial role for CAP1 in structural synaptic plasticity. By exploiting mouse hippocampal neurons, we tested this hypothesis in the present study. We found that induction of both forms of synaptic plasticity oppositely altered concentration of exogenous, myc-tagged CAP1 in dendritic spines, with chemical LTP (cLTP) decreasing and chemical LTD (cLTD) increasing it. cLTP induced spine enlargement in CAP1-deficient neurons. However, it did not increase the density of large spines, different from control neurons. cLTD induced spine retraction and spine size reduction in control neurons, but not in CAP1-KO neurons. Together, we report that postsynaptic myc-CAP1 concentration oppositely changed during cLTP and cTLD and that CAP1 inactivation modestly affected structural plasticity.

The diapause-like colorectal cancer cells induced by SMC4 attenuation are characterized by low proliferation and chemotherapy insensitivity

In response to adverse environmental conditions, embryonic development may reversibly cease, a process termed diapause. Recent reports connect this phenomenon with the non-genetic responses of tumors to chemotherapy, but the mechanisms involved are poorly understood. Here, we establish a multifarious role for SMC4 in the switching of colorectal cancer cells to a diapause-like state. SMC4 attenuation promotes the expression of three investment phase glycolysis enzymes increasing lactate production while also suppressing PGAM1. Resultant high lactate levels increase ABC transporter expression via histone lactylation, rendering tumor cells insensitive to chemotherapy. SMC4 acts as co-activator of PGAM1 transcription, and the coordinate loss of SMC4 and PGAM1 affects F-actin assembly, inducing cytokinesis failure and polyploidy, thereby inhibiting cell proliferation. These insights into the mechanisms underlying non-genetic chemotherapy resistance may have significant implications for the field, advancing our understanding of aerobic glycolysis functions in tumor and potentially informing future therapeutic strategies.

Targeting CaMKK2 inhibits actin cytoskeletal assembly to suppress cancer metastasis.

Triple-negative breast cancers (TNBCs) tend to become invasive and metastatic at early stages in their development. Despite some treatment successes in early stage localized TNBC, the rate of distant recurrence remains high, and long-term survival outcomes remain poor. In a search for new therapeutic targets for this disease, we observed that elevated expression of the serine/threonine kinase calcium/calmodulin (CaM)-dependent protein kinase kinase 2 (CaMKK2) is highly correlated with tumor invasiveness. In validation studies, genetic disruption of CaMKK2 expression or inhibition of its activity with small molecule inhibitors disrupted spontaneous metastatic outgrowth from primary tumors in murine xenograft models of TNBC. High-grade serous ovarian cancer (HGSOC), a high-risk, poor prognosis ovarian cancer subtype, shares many features with TNBC, and CaMKK2 inhibition effectively blocked metastatic progression in a validated xenograft model of this disease. Mechanistically, CaMKK2 increased the expression of the phosphodiesterase PDE1A which hydrolyzed cyclic guanosine monophosphate (cGMP) to decrease the cGMP-dependent activity of protein kinase G1 (PKG1). Inhibition of PKG1 resulted in decreased phosphorylation of vasodilator stimulated phosphoprotein (VASP), which in its hypophosphorylated state binds to and regulates F-actin assembly to facilitate cell movement. Together, these findings establish a targetable CaMKK2-PDE1A-PKG1-VASP signaling pathway that controls cancer cell motility and metastasis by impacting the actin cytoskeleton. Further, it identifies CaMKK2 as a potential therapeutic target that can be exploited to restrict tumor invasiveness in patients diagnosed with early-stage TNBC or localized HGSOC.

Young and Senescent Cells: Distinct Nuclear F-actin Patterns Upon Latrunculin B Induction

Both cellular senescence and cytoskeleton are involved in the formation of many diseases and cell signaling pathways. Although recent studies have shown that F-actin is involved in DNA damage repair, chromatin decompression, gene transcription regulation, and cell fate determination. But studies on F-actin and aging are still absence. It is unclear whether nuclear F-actin is present during cellular senescence. Here, by confocal optical sectioning and time-lapse imaging, we found acitn chrommobody-TagGFP2-NLS shows the beneficial on investigating senescent human fibroblast IMR-90 cells. To induce the nuclear F-actin assembly in single cell, we uesd Latrunculin B (latB) which a cytoplasmic F-actin polymerization inhibitor. It is currently unknown whether the nuclear F-actin cytoskeleton in young and senescent cells responds differently to latB treatment. Here, latB application induces distinct nuclear F-actin patterns and dynamics in young and senescent cells. Thus, after analyzing the results of actin dynamic we demonstrate a diverse effect of latB on the nuclear F-actin cytoskeleton in young and senescent cells.

Nuclear Actin Puncta in Senescent Cells

Both cellular senescence and the nuclear actin cytoskeleton participate in a plethora of physiological and pathological activities. However, their connection remains elusive. Recently, it is becoming increasingly apparent that filamentous actin plays important functions within the nucleus. Particularly, micron-scaled actin filaments (hereinafter referred to as F-actin) in the nucleus are of the essence in DNA damage repair, cell fate determination and age-related diseases etc. Latrunculin B (latB), a pharmaceutically applicable macrolide, can induce nuclear F-actin assembly. Now it is unclear how the nuclear actin skeleton responds to latB stimulation in senescent cells. Here, by time-lapse images, Doxo-induced senescent IMR-90 cells showed a large number of nuclear actin puncta, then the nuclear actin puncta become brighter and larger in size upon latB treatment. Some actin puncta even served as the elongation bases for the nuclear F-actin. Therefore, our results link the nuclear actin skeleton to cell aging and reveal the effect of latB treatment on the nuclear actin skeleton in aging cells.

Self-organized Rho and F-actin patterning in an artificial cell cortex.

The cell cortex, comprised of the plasma membrane and an underlying meshwork of filamentous actin (F-actin), is remodeled during a variety of essential biological processes. Previous work has shown that prior to large-scale remodeling the cell cortex is dynamically patterned with subcellular waves of F-actin assembly and disassembly, a phenomenon termed “cortical excitability”. In developing embryos, cortical excitability is generated through coupled positive and negative feedback, with rapid activation of F-actin assembly driven by the small GTPase Rho, followed by inhibition of Rho activity. These feedback loops are proposed to serve as a mechanism for amplification of active Rho signaling at the cell equator to support furrowing during cytokinesis, while also maintaining flexibility for rapid error correction. Investigating the mechanisms that support and regulate cortical patterning is currently limited by a lack of technical approaches that can bridge our understanding of biochemical feedback signaling and cortical pattern formation, including the molecular regulation of signaling molecules, membrane dynamics, and cytoskeletal remodeling. A breakthrough in this gap in knowledge has been the development of an “artificial cortex”, made from supported lipid bilayers (SLBs) and Xenopus egg extract, which successfully reconstitutes active Rho and F-actin dynamics in a cell-free system. This reconstituted system spontaneously develops two distinct types of self-organized cortical dynamics: singular excitable Rho and F-actin waves, and non-traveling oscillatory Rho and F-actin patches. Like in vivo cortical excitability, patterning in the artificial cortex depends on Rho activity and F-actin polymerization. We also find that SLB fluidity directly influences the propensity for pattern formation in the artificial cortex, which suggests dynamic membranes support cortical patterning. These findings reveal that the cell cortex is a self-organizing structure and present a novel approach for investigating mechanisms of Rho-GTPase-mediated cortical dynamics.

Regulation of actin-binding proteins by PI(4,5)P2.

The actin cytoskeleton powers membrane deformations during many cellular processes such as cell migration, morphogenesis, and endocytosis. One of the phosphoinositides phosphatidylinositol 4,5-bisphosphate [PI(4,5)P2] regulates activities of many actin-binding proteins (ABPs) including profilin, cofilin, Dia2, N-WASP, ezrin, and moesin; however, the underlying molecular mechanisms have remained elusive. Here, we applied biophysical approaches to uncover the molecular principles by which ABPs interact with phosphoinositide-containing membranes. Although ABPs bind to the membranes mainly through electrostatic interactions, we reveal that they show large differences in the affinities and dynamics of membrane interactions, and in the ranges of phosphoinositide densities that they sense. Profilin and cofilin show transient, low-affinity interactions with membranes, whereas F-actin assembly factors Dia2 and N-WASP reside on membranes for longer periods to perform their functions. Ezrin and moesin, which link the actin cytoskeleton to the plasma membrane, bind membranes with high affinities and slow dissociation dynamics. Unlike profilin, cofilin, Dia2, and N-WASP, they do not require high “stimulus-responsive” phosphoinositide density for membrane binding. Together, these findings demonstrate that membrane-interaction mechanisms of ABPs evolved to precisely perform their specific functions in cytoskeletal dynamics.

The Impact of Arp2/3 Complex Inhibition on Cytoskeleton Dynamics and Mitochondrial Function during Goat Oocyte Meiosis.

F-actin is of critical importance in oocyte meiotic maturation. Actin assembly and its dynamics are mainly regulated by actin nucleation factors. The actin-related protein complex 2/3 (Arp2/3) is responsible for the organization of F-actin filaments. However, the role of Arp2/3 complex in goat oocytes has not been fully elucidated. Our findings demonstrate that Arp2/3 complex activity is necessary for the maturation of goat oocytes. The Arp2/3 complex-specific inhibitor CK666 impairs the maturation of goat oocytes and alters the genes associated with cumulus expansion, both of which suggest that normal meiosis is affected. Arp2, one of the subunits of the Arp2/3 complex, was found to be mainly accumulated at the oocyte cortex and to co-localize with F-actin during goat oocyte maturation in our results. Thus, we further investigated the cytoskeleton dynamics and found that Arp2/3 complex inhibition disrupts the F-actin assembly and spindle organization. Further analysis revealed that, in addition to direct effects on the cytoskeleton, Arp2/3 complex could also induce ROS accumulation and oxidative stress by disrupting mitochondrial distribution and function, ultimately increasing the rate of early apoptosis in goat oocytes. Our study provides evidence that the Arp2/3 complex is a key regulator of goat oocyte maturation through its regulation of the cytoskeleton dynamics and mitochondrial function.

Metformin enhances neural precursor cells migration and functional recovery after ischemic stroke in mice

Resident neural precursor cells (NPCs) activation is a promising therapeutic strategy for brain repair. This strategy involves stimulating multiple stages of NPCs development, including proliferation, self-renewal, migration, and differentiation. Metformin, an FDA-approved diabetes drug, has been shown to promote the proliferation and differentiation of NPCs. However, it is still unclear whether metformin promotes the migration of NPCs. EVOS living cell imaging system was used for observing the migration for primary NPCs dynamically in vitro after metformin treatment. For in vivo study, a mouse model of ischemic stroke was established through middle cerebral artery occlusion (MCAO). To label the proliferating cell in subventricular zone, BrdU was injected intraperitoneally into the mice. After co-staining with BrdU and doublecortin (DCX), a marker for NPCs, the migration of Brdu and DCX double positive NPCs was detected along the rostral migratory stream (RMS) and around the infarct area using frozen brain sections. Finally, the rotarod test, corner test and beam walking were performed to evaluate the motor functions of the mice after stroke in different groups. The results showed that metformin enhanced NPCs migration in vivo and in vitro by promoting F-actin assembly and lamellipodia formation. What's more, metformin treatment also significantly reduced the infarct volume and alleviated functional dysfunction after stroke. Mechanistically, metformin promoted NPCs migration via up-regulating the CDC42 expression. Taken together, metformin represents an optimal candidate agent for neural repair that is capable of not only expanding the adult NPC population but also subsequently driving them toward the destination for neuronal differentiation.

A network of mixed actin polarity in the leading edge of spreading cells

Physical interactions of cells with the underlying extracellular matrix (ECM) play key roles in multiple cellular processes. The actin cytoskeleton is a central driver and regulator of cellular dynamics, that produces membrane-protrusions such as lamellipodia and filopodia. Here, we examined actin organization in expanding lamellipodia during early stages of cell spreading. To gain insight into the 3D actin organization, we plated fibroblasts on galectin-8 coated EM grids, an ECM protein presents in disease states. We then combined cryo-electron tomography with advanced image processing tools for reconstructing the structure of F-actin in the lamellipodia. This approach enabled us to resolve the polarity and orientation of filaments, and the structure of the Arp2/3 complexes associated with F-actin branches. We show that F-actin in lamellipodial protrusions forms a dense network with three distinct sub-domains. One consists primarily of radial filaments, with their barbed ends pointing towards the membrane, the other is enriched with parallel filaments that run between the radial fibers, in addition to an intermediate sub-domain. Surprisingly, a minor, yet significant (~10%) population of actin filaments, are oriented with their barbed-ends towards the cell center. Our results provide structural insights into F-actin assembly and dynamic reorganization in the leading edge of spreading cells.

Coordination of LMO7 with FAK Signaling Sustains Epithelial Integrity in Renal Epithelia Exposed to Osmotic Pressure

The kidney epithelial barrier has multifaceted functions in body fluids, electrolyte homeostasis, and urine production. The renal epithelial barrier (REB) frequently faces and challenges osmotic dynamics, which gives rise to osmotic pressure (a physical force). Osmotic pressure overloading can crack epithelial integrity and damage the REB. The endurance of REB to osmotic pressure forces remains obscure. LMO7 (LIM domain only 7) is a protein associated with the cell-cell junctional complex and cortical F-actin. Its upregulation was observed in cells cultured under hypertonic conditions. LMO7 is predominantly distributed in renal tubule epithelial cells. Hypertonic stimulation leads to LMO7 and F-actin assembly in the cortical stress fibers of renal epithelial cells. Hypertonic-isotonic alternation, as a pressure force pushing the plasma membrane inward/outward, was set as osmotic disturbance and was applied to test FAK signaling and LMO7 functioning in maintaining junctional integrity. LMO7 depletion in cells resulted in junctional integrity loss in the epithelial sheet-cultured hypertonic medium or hypertonic-isotonic alternation. Conversely, FAK inhibition by PF-573228 led to failure in robust cortical F-actin assembly and LMO7 association with cortical F-actin in epithelial cells responding to hypertonic stress. Epithelial integrity against osmotic stress and LMO7 and FAK signaling are involved in assembling robust cortical F-actin and maintaining junctional integrity. LMO7 elaborately manages FAK activation in renal epithelial cells, which was demonstrated excessive FAK activation present in LMO7 depleted NRK-52E cells and epithelial integrity loss when cells with LMO7 depletion were exposed to a hypertonic environment. Our data suggests that LMO7 regulates FAK activation and is responsible for maintaining REB under osmotic disturbance.

Concanavalin A inhibits human liver cancer cell migration by regulating F-actin redistribution and assembly via MAPK signaling pathway.

Concanavalin A (Con A), the first and most typical representative of the legume lectin family, has a potent anti-liver cancer effect by inducing cell apoptosis and autophagy. However, its function in human liver cancer cell migration remains unclear. The present study investigated the effect of Con A on the viability and migration of human liver cancer cells with different metastatic abilities. It was found that Con A could reduce the viability of human liver cancer cells (HCCLM3, MHCC97L and HepG2) and human hepatocytes (MIHA). In addition, Con A could inhibit human liver cancer cell migration specifically without affecting hepatocytes. Sugar inhibition assay showed that glucose-related sugar binding sites played an important role in the inhibition of Con A on human liver cancer cell migration. In addition, Con A could affect HCCLM3 migration by activating ERK1/2, JNK1/2/3 and p38 signaling in the MAPK pathway. Moreover, Con A could regulate fibrous actin (F-actin) redistribution and assembly via the MAPK signaling pathway. However, Con A had no significant effect on the activation of matrix metalloproteinase (MMP)-2 and MMP-9 in HCCLM3 cells. In conclusion, Con A may bind to glucose-related receptor protein, activating ERK1/2, JNK1/2/3 and p38 signaling in the MAPK pathway, further contracting cells and reducing the content of F-actin of single cells to inhibit HCCLM3 cell migration. These results would deepen the links between human liver cancer cell migration and related glycoproteins or signaling molecules, and may provide a new perspective for Con A as a potential anticancer agent.

Comparison of incorporation of wild type and mutated actins into sarcomeres in skeletal muscle cells: A fluorescence recovery after photobleaching study.

The α-actin mutation G15R in the nucleotide-binding pocket of skeletal muscle, causes severe actin myopathy in human skeletal muscles. Expressed in cultured embryonic quail skeletal myotubes, YFP-G15R-α-actin incorporates in sarcomeres in a pattern indistinguishable from wildtype YFP-α-actin. However, patches of YFP-G15R-α-actin form, resembling those in patients. Analyses with FRAP of incorporation of YFP-G15R-α-actin showed major differences between fast-exchanging plus ends of overlapping actin filaments in Z-bands, versus slow exchanging ends of overlapping thin filaments in the middle of sarcomeres. Wildtype skeletal muscle YFP-α-actin shows a faster rate of incorporation at plus ends of F-actin than at their minus ends. Incorporation of YFP-G15R-α-actin molecules is reduced at plus ends, increased at minus ends. The same relationship of wildtype YFP-α-actin incorporation is seen in myofibrils treated with cytochalasin-D: decreased dynamics at plus ends, increased dynamics at minus ends, and F-actin aggregates. Speculation: imbalance of normal polarized assembly of F-actin creates excess monomers that form F-actin aggregates. Two other severe skeletal muscle YFP-α-actin mutations (H40Y and V163L) not in the nucleotide pocket do not affect actin dynamics, and lack F-actin aggregates. These results indicate that normal α-actin plus and minus end dynamics are needed to maintain actin filament stability, and avoid F-actin patches.

AKT and SGK kinases regulate cell migration by altering Scar/WAVE complex activation and Arp2/3 complex recruitment.

Cell polarity and cell migration both depend on pseudopodia and lamellipodia formation. These are regulated by coordinated signaling acting through G-protein coupled receptors and kinases such as PKB/AKT and SGK, as well as the actin cytoskeletal machinery. Here we show that both Dictyostelium PKB and SGK kinases (encoded by pkbA and pkgB) are dispensable for chemotaxis towards folate. However, both are involved in the regulation of pseudopod formation and thus cell motility. Cells lacking pkbA and pkgB showed a substantial drop in cell speed. Actin polymerization is perturbed in pkbA- and reduced in pkgB- and pkbA-/pkgB- mutants. The Scar/WAVE complex, key catalyst of pseudopod formation, is recruited normally to the fronts of all mutant cells (pkbA-, pkgB- and pkbA-/pkgB-), but is unexpectedly unable to recruit the Arp2/3 complex in cells lacking SGK. Consequently, loss of SGK causes a near-complete loss of normal actin pseudopodia, though this can be rescued by overexpression of PKB. Hence both PKB and SGK are required for correct assembly of F-actin and recruitment of the Arp2/3 complex by the Scar/WAVE complex during pseudopodia formation.

Distinct functions of dimeric and monomeric scaffold protein Alix in regulating F-actin assembly and loading of exosomal cargo

Alix is a ubiquitously expressed scaffold protein that participates in numerous cellular processes related to the remodeling/repair of membranes and the actin cytoskeleton. Alix exists in monomeric and dimeric/multimeric configurations, but how dimer formation occurs and what role the dimer has in Alix-mediated processes are still largely elusive. Here, we reveal a mechanism for Alix homodimerization mediated by disulfide bonds under physiological conditions and demonstrate that the Alix dimer is enriched in exosomes and F-actin cytoskeleton subcellular fractions. Proteomic analysis of exosomes derived from Alix−/− primary cells underlined the indispensable role of Alix in loading syntenin into exosomes, thereby regulating the cellular levels of this protein. Using a set of deletion mutants, we define the function of Alix Bro1 domain, which is solely required for its exosomal localization, and that of the V domain, which is needed for recruiting syntenin into exosomes. We reveal an essential role for Cys814 within the disordered proline-rich domain for Alix dimerization. By mutating this residue, we show that Alix remains exclusively monomeric and, in this configuration, is effective in loading syntenin into exosomes. In contrast, loss of dimerization affects the ability of Alix to associate with F-actin, thereby compromising Alix-mediated cytoskeleton remodeling. We propose that dimeric and monomeric forms of Alix selectively execute two of the protein’s main functions: exosomal cargo loading and cytoskeleton remodeling.

Protective effect of resveratrol against cadmium-induced toxicity on ovine oocyte in vitro maturation and fertilization

BackgroundHeavy metal cadmium (Cd) is a widespread environmental contaminant with a potential toxicity that might negatively affect female reproduction and fertility. It has been reported that Cd exposure impaired the quality of oocytes and led to a defective maturation and fertilization, through oxidative stress induction. Resveratrol (Res) is a natural polyphenol with strong antioxidant properties that exhibited protective role in preventing oocyte redox homeostasis disruption and quality decline. Here, we explored whether the addition of Res to in vitro maturation (IVM) medium might act as a protection against Cd-induced toxicity on ovine oocyte maturation and fertilization. Firstly, we evaluated the effect of supplementing IVM medium with two different Res concentrations (1 and 2 μmol/L) on nuclear maturation and fertilization of oocytes matured under CdCl2 (2 μmol/L) exposure. Therefore, the concentration of 1 μmol/L Res was selected to analyse the effects of this compound on intracellular ROS levels, mitochondrial (mt) distribution and activity, chromatin configuration, cytoskeleton morphology, cortical granules (CGs) distribution and mRNA expression of genes associated with cellular response to oxidative stress (i.e. SIRT1, SOD 1, GPX1, GSR, CAT) in Cd-exposed in vitro matured oocytes.ResultsWe found that 1 μmol/L Res restored the reduced oocyte meiotic competence induced by Cd exposure as well as, Res sustained oocyte ability to be normally fertilized and decreased polyspermic fertilization at both tested concentrations. Moreover, we demonstrated that 1 μmol/L Res mitigated Cd-induced alterations of oocyte cytoplasmic maturation by reducing reactive oxygen species (ROS) accumulation, preventing mt dysfunction, maintaining the correct meiotic spindle and cortical F-actin assembly and the normal cortical granule distribution as well as up-regulating SIRT1, SOD1 and GPX1 genes.ConclusionsTaken together, our findings highlighted the beneficial influence exerted by Res in preventing Cd-induced disturbance of nuclear and cytoplasmic maturation and subsequent fertilization in ovine oocytes. Res treatment may help to establish defence strategies counteracting Cd-induced toxicity on the female gamete.

Keratin5-cytoskeleton-BMP4 network regulates cell phenotype conversions during cardiac regeneration

The reconstruction of a blood supply system and myocardial recovery from inflamamtory reactions in the infract zone remains a challenge in cardiac regeneration after myocardial infarction. Here, we observed that the local myocardial cells and the clotted blood cells undergo cellular remodeling via cytoplasmic exocytosis and nuclear reorganization in zebrafish hearts after resection of the ventricular apex. The subsequent tissue regeneration processes were visualized by detection of the spatiotemporal expression of three tissue specific genes (α-SMA which marks for vasculature/fibrogenesis, Flk1for angiogenesis/hematopoiesis, and Pax3a for remusculogensis), and two histone modification markers (H3K9Ac and H3K9Me3 for chromatin remodeling). By analyzing the composition of the blastema tissue fractions we found that Krt5 peptide could promote F-actin assembly, BMP4-pSmad2/5/8 signaling activity, and H3K9Me3-mediated chromatin accessibility at the blastema representative genes in the cultured zebrafish embryonic fibroblasts. Further in vivo tests demonstrated that Krt5 interacted with beta actin, and promoted Gata3 expression and Flk1-GFP marked blastema angiogenesis. These results proposed a new Krt5-cytoskeleton-BMP4 mechanotransduction mechanism in the epithelial-dependent and cell phenotype conversion-based tissue regeneration.

MEOX2-mediated regulation of Cathepsin S promotes cell proliferation and motility in glioma

Nuclear transcription factor Mesenchyme Homeobox 2 (MEOX2) is a homeobox gene that is originally discovered to suppress the growth of vascular smooth muscle and endothelial cells. However, whether or not it is connected to cancer is yet unknown. Here, we report that MEOX2 functions as a tumor-initiating element in glioma. Bioinformatic analyses of public databases and investigation of MEOX2 expression in patients with glioma demonstrated that MEOX2 was abundant at both mRNA and protein levels in glioma. MEOX2 expression was shown to be inversely linked with the prognosis of glioma patients. MEOX2 inhibition changed the morphology of glioma cells, inhibited cell proliferation and motility, whereas had no effect on cell apoptosis. Besides, silencing MEOX2 also hampered the epithelial-mesenchymal transition (EMT), focal adhesion formation, and F-actin assembly. Overexpression of MEOX2 exhibited opposite effects. Importantly, RNA-sequencing, ChIP-qPCR assay, and luciferase reporter assay revealed Cathepsin S (CTSS) as a novel transcriptional target of MEOX2 in glioma cells. Consistently, MEOX2 causes glioma tumor development in mice and greatly lowers the survival period of tumor-bearing mice. Our findings indicate that MEOX2 promotes tumorigenesis and progression of glioma partially through the regulation of CTSS. Targeting MEOX2-CTSS axis might be a promising alternative for the treatment of glioma.

The prophase oocyte nucleus is a homeostatic G-actin buffer.

ABSTRACTFormation of healthy mammalian eggs from oocytes requires specialised F-actin structures. F-actin disruption produces aneuploid eggs, which are a leading cause of human embryo deaths, genetic disorders and infertility. We found that oocytes contain prominent nuclear F-actin structures that are correlated with meiotic developmental capacity. We demonstrate that nuclear F-actin is a conserved feature of healthy mammalian oocytes and declines significantly with female reproductive ageing. Actin monomers used for nuclear F-actin assembly are sourced from an excess pool in the oocyte cytoplasm. Increasing monomeric G-actin transfer from the cytoplasm to the nucleus or directly enriching the nucleus with monomers led to assembly of stable nuclear F-actin bundles that significantly restrict chromatin mobility. By contrast, reducing G-actin monomer transfer by blocking nuclear import triggered assembly of a dense cytoplasmic F-actin network that is incompatible with healthy oocyte development. Overall, our data suggest that the large oocyte nucleus helps to maintain cytoplasmic F-actin organisation and that defects in this function are linked with reproductive age-related female infertility.This article has an associated First Person interview with Federica Giannini, joint first author of the paper.

Multiple Roles of Actin in Exo- and Endocytosis.

Cytoskeletal filamentous actin (F-actin) has long been considered a molecule that may regulate exo- and endocytosis. However, its exact roles remained elusive. Recent studies shed new light on many crucial roles of F-actin in regulating exo- and endocytosis. Here, this progress is reviewed from studies of secretory cells, particularly neurons and endocrine cells. These studies reveal that F-actin is involved in mediating all kinetically distinguishable forms of endocytosis, including ultrafast, fast, slow, bulk, and overshoot endocytosis, likely via membrane pit formation. F-actin promotes vesicle replenishment to the readily releasable pool most likely via active zone clearance, which may sustain synaptic transmission and overcome short-term depression of synaptic transmission during repetitive firing. By enhancing plasma membrane tension, F-actin promotes fusion pore expansion, vesicular content release, and a fusion mode called shrink fusion involving fusing vesicle shrinking. Not only F-actin, but also the F-actin assembly pathway, including ATP hydrolysis, N-WASH, and formin, are involved in mediating these roles of exo- and endocytosis. Neurological disorders, including spinocerebellar ataxia 13 caused by Kv3.3 channel mutation, may involve impairment of F-actin and its assembly pathway, leading in turn to impairment of exo- and endocytosis at synapses that may contribute to neurological disorders.