Membrane insertion of soluble CLIC1 into active chloride channels is triggered by specific divalent cations

The CLIC family of proteins display the unique feature of altering their structure from a soluble form to a membrane-bound chloride channel. CLIC1, a member of this family, can be found in the cytoplasm or in nuclear, ER and plasma membranes, with membrane overexpression linked to tumour proliferation. The molecular switch promoting CLIC1 membrane insertion has been related to environmental factors, but still remains unclear. Here, we use solution NMR studies to confirm that both the soluble and membrane bound forms are in the same oxidation state. Our data from fluorescence assays and chloride efflux assays indicate that Ca2+ and Zn2+ trigger association to the membrane into active chloride channels. We use fluorescence microscopy to confirm that an increase of the intracellular Ca2+ leads to re-localisation of CLIC1 to both plasma and internal membranes. Finally, we show that soluble CLIC1 adopts an equilibrium of oligomeric species, and Ca2+/Zn2+ mediated membrane insertion promotes the formation of a tetrameric assembly. Thus, our results identify Ca2+ and Zn2+ binding as the molecular switch promoting CLIC1 membrane insertion.SIGNIFICANCE STATEMENT CLIC1, a member of the CLIC family of proteins, is expressed as a soluble protein in cells but can insert in the membrane forming a chloride channel. This chloride channel form is upregulated in different types of cancers including glioblastoma and promote tumour invasiveness and metastasis. The factors promoting CLIC1 membrane insertion nor the mechanism of this process are yet understood. Here, we use a combination of solution NMR, biophysics and fluorescence microscopy to identify Ca2+ and Zn2+ binding as the switch to promote CLIC1 insertion into the membrane to form active chloride channels. We also provide a simple mechanism how such transition to the membrane occurs. Such understanding will enable subsequent studies on the structure of the chloride channel form and its inhibition

Evaluación de la capacidad antioxidante y estabilidad térmica de la cáscara, zumo y semilla de lima dulce, limón rugoso y limón tipo mandarina

El objetivo de la investigación fue evaluar la capacidad antioxidante y la estabilidad térmica en la cáscara, zumo y semilla de lima dulce, limón rugoso y limón tipo mandarina. Se determinó el contenido de fenoles totales, capacidad antioxidante en función a los radicales DPPH y ABTS, expresados como equivalentes de Trolox. El contenido de fenoles totales en cáscara, semilla y zumo, de lima dulce fue 1,08±0,00 g EAG/100mL, 0,16±0,00 g EAG/100g y 0,29±0,00 g EAG/100g; en limón rugoso fue 0,97±0,01 g EAG/100mL, 0,19±0,00 g EAG/100g y 0,09±0,00 g EAG/100g y en limón mandarina fue 0,83±0,01 g EAG/100mL, 0,15±0,01 g EAG/100g y 0,08±0,00 g EAG/100g. No se detectó la presencia de antocianinas. Se logró determinar la capacidad antioxidante en función a los radicales DPPH y ABTS, expresados como Trolox equivalente, la capacidad antioxidante en cáscaras vario desde 71,87±0,03 a 730,75±0,03 uMTEAC/100g b.s. en DPPHº+ y de 63,73±1,22 a 78,18±0,58 uMTEAC/g b.s. en ABTSº+; en semillas desde 200,72±0,02 a 257,45±0,06 uMTEAC/100g b.s. en DPPHº+ y de 8,45±0,20 a 12,12±0,19 uMTEAC/g b.s. en ABTSº+ y en zumo de 335,60±9,00 a 602,23±,08uMTEAC/mL b.s. en DPPHº+ y de 61,35±3,66 a 301,47±39,87uMTEAC/100mL b.s. en ABTSº+. Los fenoles totales y capacidad antioxidante como radical DPPH, en cáscara y semillas de limón rugoso y limón mandarina, permanecieron termoestables, durante el almacenamiento; la capacidad antioxidante expresada como radical ABTS, disminuyo. Los polifenoles y capacidad antioxidante expresados como radicales DPPH y ABTS, en el jugo de limón dulce, mostraron un incremento durante el almacenamiento a diversas temperaturas

Azetidinium lead iodide for perovskite solar cells

Hybrid organic–inorganic perovskites have been established as good candidate materials for emerging photovoltaics, with device efficiencies of over 22% being reported. There are currently only two organic cations, methylammonium and formamidinium, which produce 3D perovskites with band gaps suitable for photovoltaic devices. Numerous computational studies have identified azetidinium as a potential third cation for synthesizing organic–inorganic perovskites, but to date no experimental reports of azetidinium containing perovskites have been published. Here we prepare azetidinium lead iodide for the first time. Azetidinium lead iodide is a stable, bright orange material which does not appear to form a 3D or a 2D perovskite. It was successfully used as the absorber layer in solar cells. We also show that it is possible to make mixed cation devices by adding the azetidinium cation to methylammonium lead iodide. Computational studies show that the substitution of up to 5% azetidinium into the methylammonium lead iodide is energetically favourable and that phase separation does not occur at these concentrations. Mixed azetidinium–methylammonium cells show improved performance and reduced hysteresis compared to methylammonium lead iodide cells

Understanding the role of Cesium and Rubidium additives in perovskite solar cells: trap states, charge transport, and recombination

Adding cesium (Cs) and rubidium (Rb) cations to FA0.83MA0.17Pb(I0.83Br0.17)3 hybrid lead halide perovskites results in a remarkable improvement in solar cell performance, but the origin of the enhancement has not been fully understood yet. In this work, time‐of‐flight, time‐resolved microwave conductivity, and thermally stimulated current measurements are performed to elucidate the impact of the inorganic cation additives on the trap landscape and charge transport properties within perovskite solar cells. These complementary techniques allow for the assessment of both local features within the perovskite crystals and macroscopic properties of films and full devices. Strikingly, Cs‐incorporation is shown to reduce the trap density and charge recombination rates in the perovskite layer. This is consistent with the significant improvements in the open‐circuit voltage and fill factor of Cs‐containing devices. By comparison, Rb‐addition results in an increased charge carrier mobility, which is accompanied by a minor increase in device efficiency and reduced current–voltage hysteresis. By mixing Cs and Rb in quadruple cation (Cs‐Rb‐FA‐MA) perovskites, the advantages of both inorganic cations can be combined. This study provides valuable insights into the role of these additives in multiple‐cation perovskite solar cells, which are essential for the design of high‐performance devices

Migration of cations induces reversible performance losses over day/night cycling in perovskite solar cells

Perovskites have been demonstrated in solar cells with a power conversion efficiency of well above 20​%, which makes them one of the strongest contenders for next generation photovoltaics. While there are no concerns about their efficiency, very little is known about their stability under illumination and load. Ionic defects and their migration in the perovskite crystal lattice are some of the most alarming sources of degrdn., which can potentially prevent the commercialization of perovskite solar cells (PSCs)​. In this work, we provide direct evidence of elec. field-​induced ionic defect migration and we isolate their effect on the long-​term performance of state-​of-​the-​art devices. Supported by modeling, we demonstrate that ionic defects, migrating on timescales significantly longer (above 103 s) than what has so far been explored (from 10-​1 to 102 s)​, abate the initial efficiency by 10-​15​% after several hours of operation at the max. power point. Though these losses are not negligible, we prove that the initial efficiency is fully recovered when leaving the device in the dark for a comparable amt. of time. We verified this behavior over several cycles resembling day​/night phases, thus probing the stability of PSCs under native working conditions. This unusual behavior reveals that research and industrial stds. currently in use to assess the performance and the stability of solar cells need to be adjusted for PSCs. Our work paves the way for much needed new testing protocols and figures of merit specifically designed for PSCs

Structure-based discovery and in vitro validation of inhibitors of chloride intracellular channel 4 protein

The use of computer-aided methods have continued to propel accelerated drug discovery across various disease models, interestingly allowing the specific inhibition of pathogenic targets. Chloride Intracellular Channel Protein 4 (CLIC4) is a novel class of intracellular ion channel highly implicated in tumor and vascular biology. It regulates cell proliferation, apoptosis and angiogenesis; and is involved in multiple pathologic signaling pathways. Absence of specific inhibitors however impedes its advancement to translational research. Here, we integrate structural bioinformatics and experimental research approaches for the discovery and validation of small-molecule inhibitors of CLIC4. High-affinity allosteric binders were identified from a library of 1615 Food and Drug Administration (FDA)-approved drugs via a high-performance computing-powered blind-docking approach, resulting in the selection of amphotericin B and rapamycin. NMR assays confirmed the binding and conformational disruptive effects of both drugs while they also reversed stress-induced membrane translocation of CLIC4 and inhibited endothelial cell migration. Structural and dynamics simulation studies further revealed that the inhibitory mechanisms of these compounds were hinged on the allosteric modulation of the catalytic glutathione (GSH)-like site loop and the extended catalytic β loop which may elicit interference with the catalytic activities of CLIC4. Structure-based insights from this study provide the basis for the selective targeting of CLIC4 to treat the associated pathologies

Organic gelators as growth control agents for stable and reproducible hybrid perovskite-based solar cells

Low molecular-weight organic gelators are widely used to influence the solidification of polymers, with applications ranging from packaging items, food containers to organic electronic devices, including organic photovoltaics. Here, this concept is extended to hybrid halide perovskite-based materials. In-situ time-resolved grazing incidence wide angle x-ray scattering (GIWAXS) measurements performed during spin-coating reveal that organic gelators beneficially influence the nucleation and growth of the perovskite precursor phase. This can be exploited for the fabrication of planar n-i-p heterojunction devices with MAPbI3 (MA = CH3NH3 +) that display a performance that not only is enhanced by ~25% compared to solar cells where the active layer was produced without the use of a gelator but that also feature a higher stability to moisture and a reduced hysteresis. Most importantly, the presented approach is straight-forward and simple, and it provides a general method to render the film-formation of hybrid perovskites more reliable and robust, analogous to the control that is afforded by these additives in the processing of commodity ‘plastics’

A quantitative assay to study the lipid selectivity of membrane-associated systems using solution NMR

The activity of membrane proteins and compounds that interact with the membrane is modulated by the surrounding lipid composition. However, there are no simple methods that determine the composition of these annular phospholipids in eukaryotic systems. Herein, we describe a simple methodology that enables the identification and quantification of the lipid composition around membrane-associated compounds using SMA-nanodiscs and routine $^1H-^{31}P NMR$

Time-Dependent COVID-19 Mortality in Patients with Cancer: An Updated Analysis of the OnCovid Registry

Importance: Whether the severity and mortality of COVID-19 in patients with cancer have improved in terms of disease management and capacity is yet to be defined. Objective: To test whether severity and mortality from COVID-19 among patients with cancer have improved during the course of the pandemic. Design, Setting, and Participants: OnCovid is a European registry that collects data on consecutive patients with solid or hematologic cancer and COVID-19. This multicenter case series study included real-world data from 35 institutions across 6 countries (UK, Italy, Spain, France, Belgium, and Germany). This update included patients diagnosed between February 27, 2020, and February, 14, 2021. Inclusion criteria were confirmed diagnosis of SARS-CoV-2 infection and a history of solid or hematologic cancer. Exposures: SARS-CoV-2 infection. Main Outcomes and Measures: Deaths were differentiated at 14 days and 3 months as the 2 landmark end points. Patient characteristics and outcomes were compared by stratifying patients across 5 phases (February to March 2020, April to June 2020, July to September 2020, October to December 2020, and January to February 2021) and across 2 major outbreaks (February to June 2020 and July 2020 to February 2021). Results: At data cutoff, 2795 consecutive patients were included, with 2634 patients eligible for analysis (median [IQR] age, 68 [18-77] years; 52.8% men). Eligible patients demonstrated significant time-dependent improvement in 14-day case-fatality rate (CFR) with estimates of 29.8% (95% CI, 0.26-0.33) for February to March 2020; 20.3% (95% CI, 0.17-0.23) for April to June 2020; 12.5% (95% CI, 0.06-22.90) for July to September 2020; 17.2% (95% CI, 0.15-0.21) for October to December 2020; and 14.5% (95% CI, 0.09-0.21) for January to February 2021 (all P <.001) across the predefined phases. Compared with the second major outbreak, patients diagnosed in the first outbreak were more likely to be 65 years or older (974 of 1626 [60.3%] vs 564 of 1008 [56.1%]; P =.03), have at least 2 comorbidities (793 of 1626 [48.8%] vs 427 of 1008 [42.4%]; P =.001), and have advanced tumors (708 of 1626 [46.4%] vs 536 of 1008 [56.1%]; P <.001). Complications of COVID-19 were more likely to be seen (738 of 1626 [45.4%] vs 342 of 1008 [33.9%]; P <.001) and require hospitalization (969 of 1626 [59.8%] vs 418 of 1008 [42.1%]; P <.001) and anti-COVID-19 therapy (1004 of 1626 [61.7%] vs 501 of 1008 [49.7%]; P <.001) during the first major outbreak. The 14-day CFRs for the first and second major outbreaks were 25.6% (95% CI, 0.23-0.28) vs 16.2% (95% CI, 0.13-0.19; P <.001), respectively. After adjusting for country, sex, age, comorbidities, tumor stage and status, anti-COVID-19 and anticancer therapy, and COVID-19 complications, patients diagnosed in the first outbreak had an increased risk of death at 14 days (hazard ratio [HR], 1.85; 95% CI, 1.47-2.32) and 3 months (HR, 1.28; 95% CI, 1.08-1.51) compared with those diagnosed in the second outbreak. Conclusions and Relevance: The findings of this registry-based study suggest that mortality in patients with cancer diagnosed with COVID-19 has improved in Europe; this improvement may be associated with earlier diagnosis, improved management, and dynamic changes in community transmission over time.