FG/FxFG as well as GLFG repeats form a selective permeability barrier with self-healing properties.
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ABSTRACT:
SUBMITTER: Frey S
PROVIDER: S-EPMC2728434 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Nup98 FG repeat domains comprise hydrophobic FG motifs linked through uncharged spacers. FG motifs capture nuclear transport receptors (NTRs) during nuclear pore complex (NPC) passage, confer inter-repeat cohesion, and condense the domains into a selective phase with NPC-typical barrier properties. We show that shortening inter-FG spacers enhances cohesion, increases phase density, and tightens such barrier - all consistent with a sieve-like phase. Phase separation tolerates mutating the Nup98-typical GLFG motifs, provided domain-hydrophobicity remains preserved. NTR-entry, however, is sensitive to (certain) deviations from canonical FG motifs, suggesting co-evolutionary adaptation. Unexpectedly, we observed that arginines promote FG-phase-entry apparently also by hydrophobic interactions/ hydrogen-bonding and not just through cation-π interactions. Although incompatible with NTR·cargo complexes, a YG phase displays remarkable transport selectivity, particularly for engineered GFPNTR-variants. GLFG to FSFG mutations make the FG phase hypercohesive, precluding NTR-entry. Extending spacers relaxes this hypercohesion. Thus, antagonism between cohesion and NTR·FG interactions is key to transport selectivity.
| S-EPMC9918544 | biostudies-literature
Project description:Low-viscosity oils could potentially act as self-healing barrier coatings because they can readily flow and reconnect to heal minor damage. For the same reason, however, they typically do not form stable coatings on metal surfaces. Increasing viscosity helps to stabilize the oil coating, but it also slows down the healing process. Here, we report a strategy for creating highly stable oil coatings on metal surfaces without sacrificing their remarkable self-healing properties. Low-viscosity oil films can be immobilized on metal surfaces using lightweight microcapsules as thickeners, which form a dynamic network to prevent the creep of the coating. When the coating is scratched, oil around the opening can rapidly flow to cover the exposed area, reconnecting the particle network. Use of these coatings as anticorrosion barriers is demonstrated. The coatings can be easily applied on metal surfaces, including those with complex geometries, both in air or under water, and remain stable even in turbulent water. They can protect metal in corrosive environments for extended periods of time and can self-heal repeatedly when scratched at the same spot. Such a strategy may offer effective mitigation of the dangerous localized corrosion aggravated by minor imperfections or damage in protective coatings, which are typically hard to prevent or detect, but can drastically degrade metal properties.
| S-EPMC6750121 | biostudies-literature
Project description:Nuclear Pore Complexes (NPCs) regulate bidirectional transport between the nucleus and the cytoplasm. Intrinsically disordered FG-Nups line the NPC lumen and form a selective barrier, where transport of most proteins is inhibited whereas specific transporter proteins freely pass. The mechanism underlying selective transport through the NPC is still debated. Here, we reconstitute the selective behaviour of the NPC bottom-up by introducing a rationally designed artificial FG-Nup that mimics natural Nups. Using QCM-D, we measure selective binding of the artificial FG-Nup brushes to the transport receptor Kap95 over cytosolic proteins such as BSA. Solid-state nanopores with the artificial FG-Nups lining their inner walls support fast translocation of Kap95 while blocking BSA, thus demonstrating selectivity. Coarse-grained molecular dynamics simulations highlight the formation of a selective meshwork with densities comparable to native NPCs. Our findings show that simple design rules can recapitulate the selective behaviour of native FG-Nups and demonstrate that no specific spacer sequence nor a spatial segregation of different FG-motif types are needed to create selective NPCs.
| S-EPMC8012357 | biostudies-literature
Project description:Efficient delivery of therapeutics across the neuroprotective blood-brain barrier (BBB) remains a formidable challenge for central nervous system drug development. High-fidelity in vitro models of the BBB could facilitate effective early screening of drug candidates targeting the brain. In this study, we developed a microfluidic BBB model that is capable of mimicking in vivo BBB characteristics for a prolonged period and allows for reliable in vitro drug permeability studies under recirculating perfusion. We derived brain microvascular endothelial cells (BMECs) from human induced pluripotent stem cells (hiPSCs) and cocultured them with rat primary astrocytes on the two sides of a porous membrane on a pumpless microfluidic platform for up to 10 days. The microfluidic system was designed based on the blood residence time in human brain tissues, allowing for medium recirculation at physiologically relevant perfusion rates with no pumps or external tubing, meanwhile minimizing wall shear stress to test whether shear stress is required for in vivo-like barrier properties in a microfluidic BBB model. This BBB-on-a-chip model achieved significant barrier integrity as evident by continuous tight junction formation and in vivo-like values of trans-endothelial electrical resistance (TEER). The TEER levels peaked above 4000 Ω · cm2 on day 3 on chip and were sustained above 2000 Ω · cm2 up to 10 days, which are the highest sustained TEER values reported in a microfluidic model. We evaluated the capacity of our microfluidic BBB model to be used for drug permeability studies using large molecules (FITC-dextrans) and model drugs (caffeine, cimetidine, and doxorubicin). Our analyses demonstrated that the permeability coefficients measured using our model were comparable to in vivo values. Our BBB-on-a-chip model closely mimics physiological BBB barrier functions and will be a valuable tool for screening of drug candidates. The residence time-based design of a microfluidic platform will enable integration with other organ modules to simulate multi-organ interactions on drug response. Biotechnol. Bioeng. 2017;114: 184-194. © 2016 Wiley Periodicals, Inc.
| S-EPMC6650146 | biostudies-literature
Project description:Conformational changes at supramolecular interfaces are fundamentally coupled to binding activity, yet it remains a challenge to probe this relationship directly. Within the nuclear pore complex, this underlies how transport receptors known as karyopherins proceed through a tethered layer of intrinsically disordered nucleoporin domains containing Phe-Gly (FG)-rich repeats (FG domains) that otherwise hinder passive transport. Here, we use nonspecific proteins (i.e., BSA) as innate molecular probes to explore FG domain conformational changes by surface plasmon resonance. This mathematically diminishes the surface plasmon resonance refractive index constraint, thereby providing the means to acquire and correlate height changes in a surface-tethered FG domain layer to Kap binding affinities in situ with respect to their relative spatial arrangements. Stepwise measurements show that FG domain collapse is caused by karyopherin ?1 (Kap?1) binding at low concentrations, but this gradually transitions into a reextension at higher Kap?1 concentrations. This ability to self-heal is intimately coupled to Kap?1-FG binding avidity that promotes the maximal incorporation of Kap?1 into the FG domain layer. Further increasing Kap?1 to physiological concentrations leads to a "pileup" of Kap?1 molecules that bind weakly to unoccupied FG repeats at the top of the layer. Therefore, binding avidity does not hinder fast transport per se. Revealing the biophysical basis underlying the form-function relationship of Kap?1-FG domain behavior results in a convergent picture in which transport and mechanistic aspects of nuclear pore complex functionality are reconciled.
| S-EPMC3479521 | biostudies-other
Project description:Nuclear pore complexes (NPCs) are ∼100 MDa transport channels assembled from multiple copies of ∼30 nucleoporins (Nups). One-third of these Nups contain phenylalanine-glycine (FG)-rich repeats, forming a diffusion barrier, which is selectively permeable for nuclear transport receptors that interact with these repeats. Here, we identify an additional function of FG repeats in the structure and biogenesis of the yeast NPC. We demonstrate that GLFG-containing FG repeats directly bind to multiple scaffold Nups in vitro and act as NPC-targeting determinants in vivo. Furthermore, we show that the GLFG repeats of Nup116 function in a redundant manner with Nup188, a nonessential scaffold Nup, to stabilize critical interactions within the NPC scaffold needed for late steps of NPC assembly. Our results reveal a previously unanticipated structural role for natively unfolded GLFG repeats as Velcro to link NPC subcomplexes and thus add a new layer of connections to current models of the NPC architecture.
| S-EPMC5992322 | biostudies-literature
Project description:Vascular endothelial growth factor (VEGF) contributes to blood-retinal barrier (BRB) dysfunction in several blinding eye diseases, including diabetic retinopathy. Signaling via the secreted protein norrin through the frizzled class receptor 4 (FZD4)/LDL receptor-related protein 5-6 (LRP5-6)/tetraspanin 12 (TSPAN12) receptor complex is required for developmental vascularization and BRB formation. Here, we tested the hypothesis that norrin restores BRB properties after VEGF-induced vascular permeability in diabetic rats or in animals intravitreally injected with cytokines. Intravitreal co-injection of norrin with VEGF completely ablated VEGF-induced BRB permeability to Evans Blue-albumin. Likewise, 5-month diabetic rats exhibited increased permeability of FITC-albumin, and a single norrin injection restored BRB properties. These results were corroborated in vitro, where co-stimulation of norrin with VEGF or stimulation of norrin after VEGF exposure restored barrier properties, indicated by electrical resistance or 70-kDa RITC-dextran permeability in primary endothelial cell culture. Interestingly, VEGF promoted norrin signaling by increasing the FZD4 co-receptor TSPAN12 at cell membranes in an MAPK/ERK kinase (MEK)/ERK-dependent manner. Norrin signaling through β-catenin was required for BRB restoration, but glycogen synthase kinase 3 α/β (GSK-3α/β) inhibition did not restore BRB properties. Moreover, levels of the tight junction protein claudin-5 were increased with norrin and VEGF or with VEGF alone, but both norrin and VEGF were required for enriched claudin-5 localization at the tight junction. These results suggest that VEGF simultaneously induces vascular permeability and promotes responsiveness to norrin. Norrin, in turn, restores tight junction complex organization and BRB properties in a β-catenin-dependent manner.
| S-EPMC7135996 | biostudies-literature
Project description:This paper investigates the effectiveness of a specific crystalline waterproofing admixture (CWA) in concrete as a function of a water-binder ratio. Four concrete mixes with and without CWA were prepared; two of them with a water-binder ratio of 0.45 and two of them with a water-binder ratio of 0.55. Water permeability and compressive strength were tested on hardened concrete specimens and self-healing of cracks over time was observed. Cement paste and CWA paste were prepared to clarify the results obtained on the concrete specimens. SEM and EDS and XRD and FTIR were performed on the hardened pastes to explain the mechanism of CWA working. The results show that the addition of CWA had no significant effect on the compressive strength of the concrete, but reduced the water penetration depth in the concrete, and the reduction was more effective for mixes with lower water-binder ratio. Regarding the self-healing effect, it can be concluded that the addition of CWA improves the crack healing in concrete, but the efficiency of self-healing is highly dependent on the initial crack width. The mechanisms involved in the reduction of water penetration depth and crack healing in concrete can be explained by different mechanisms; one is creation of the CSH gel from unreacted clinker grains, then formation carbonate, and additional mechanism is gel formation (highly expansive Mg-rich hydro-carbonate) from magnesium based additives. The presence of sodium silicate, which would transform into carbonate/bicarbonate, also cannot be excluded.
| S-EPMC8069652 | biostudies-literature
Project description:Rationale. The microvasculature of the central nervous system includes the blood-brain barrier (BBB), which regulates the permeability to nutrients and restricts the passage of toxic agents and inflammatory cells. Canonical Wnt/b-catenin signaling is responsible for the early phases of brain vascularization and blood-brain barrier differentiation. However, this signal declines after birth and other signaling pathways able to maintain barrier integrity at postnatal stage are still unknown. Objective. Sox17 constitutes a major downstream target of Wnt/b-catenin in endothelial cells and regulates arterial differentiation. In the present paper, we asked whether Sox17 may act downstream of Wnt/b-catenin in inducing BBB differentiation and maintenance. Methods and Results. Using reporter mice and nuclear staining of Sox17 and b-catenin, we report that while b-catenin signaling declines after birth, Sox17 activation increases and remains high in the adult. Endothelial-specific inactivation of Sox17 leads to increase of permeability of the brain microcirculation. The severity of this effect depends on the degree of BBB maturation: it is strong in the embryo, and progressively declines after birth. In search of Sox17 mechanism of action, RNA-Seq analysis of gene expression of brain endothelial cells has identified members of the Wnt/b-catenin signaling pathway as downstream targets of Sox17. Consistently, we found that Sox17 is a positive inducer of Wnt/b-catenin signaling and it acts in concert with this pathway to induce and maintain BBB properties. In vivo, inhibition of the b-catenin destruction complex or expression of a degradation-resistant b-catenin mutant, prevent the increase in permeability and retina vascular malformations observed in the absence of Sox17. Conclusions. Our data highlight a novel role for Sox17 in the induction and maintenance of the BBB and they underline the strict reciprocal tuning of this transcription factor and Wnt/b-catenin pathway. Modulation of Sox17 activity may be relevant to control BBB permeability in pathological conditions.
2019-02-12 | GSE122564 | GEO
Project description:The effects of the soft block fraction and H-bond state in thermoplastic polyurethanes on autonomous entropy-driven scratch closure and barrier restoration are studied. To this aim, comparable polyurethanes with different segmentation states are applied as organic coatings on plain carbon steel plates, scratched under very well-controlled conditions, and the scratch closure and sealing kinetics are studied in detail. The scratch closure is measured optically, while the barrier restoration is probed by the accelerated cyclic electrochemical technique (ACET). Scratch closure, attributed to entropic elastic recovery (EER), is followed in a marked two-step process by barrier restoration governed by local viscous flow and the state of the interfacial hydrogen bonding. Polyurethanes with a lower soft phase fraction lead to a higher urea/urethane ratio, which in turn influences the healing efficiency of each healing step. Interestingly, softer polyurethanes leading to efficient crack closure were unable to sufficiently restore barrier properties. The present work highlights the critical role of the soft/hard block and urea/urethane H-bond state content on crack closure and barrier restoration of anticorrosive organic coatings and points at design rules for the design of more efficient corrosion-protective self-healing polyurethanes.
| S-EPMC8154210 | biostudies-literature