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SUMMARYIntraflagellar transport (IFT) trains, built around IFT-A and IFT-B complexes, are carried by opposing motors to import and export ciliary cargo. While transported by kinesin-2 on anterograde IFT trains, the dynein-2 motor adopts an autoinhibitory conformation until it needs to be activated at the ciliary tip to power retrograde IFT. Growing evidence has linked the IFT-A complex to retrograde IFT; however, its roles in this process remain unknown.Here, we used CRISPR/Cas9-mediated editing to disable the dynein-2 autoinhibition mechanism inCaenorhabditis elegans, and assessed its impact on IFT with high-resolution live imaging and photobleaching analyses. Remarkably, this dynein-2 “hot-wiring” approach reignites retrograde motility inside IFT-A-deficient cilia, without triggering tug-of-war events. In addition to providing functional evidence that multiple mechanisms maintain dynein-2 inhibited during anterograde IFT, our data uncover key roles for IFT-A in: mediating motor-train coupling during IFT turnaround; promoting retrograde IFT initiation; and modulating dynein-2 retrograde motility.Highlights- Hot-wiring mutations enable dynein-2 to undergo retrograde movement in IFT-A-deficient cilia- Disabling dynein-2 autoinhibition reveals that multiple mechanisms restrain dynein-2 activity during anterograde IFT- IFT-A promotes retrograde IFT initiation and efficient dynein-2 motility- IFT-A mediates dynein-2 coupling to retrograde IFT trains

The dynein-2 motor complex drives retrograde intraflagellar transport (IFT), playing a pivotal role in the assembly and functions of cilia. However, the mechanisms that regulate dynein-2 motility remain poorly understood. Here, we identify the Caenorhabditis elegans WDR60 homologue, WDR-60, and dissect the roles of this intermediate chain using genome editing and live imaging of endogenous dynein-2/IFT components. We find that loss of WDR-60 impairs dynein-2 recruitment to cilia and its incorporation onto anterograde IFT trains, reducing retrograde motor availability at the ciliary tip. Consistent with this, we show that fewer dynein-2 motors power WDR-60-deficient retrograde IFT trains, which move at reduced velocities and fail to exit cilia, accumulating on the distal side of the transition zone. Remarkably, disrupting the transition zone's NPHP module almost fully restores ciliary exit of underpowered retrograde trains in wdr-60 mutants. This work establishes WDR-60 as a major contributor to IFT, and the NPHP module as a roadblock to dynein-2 passage through the transition zone. This work was financed by Fundo Europeu de Desenvolvimento Regional (FEDER) through the COMPETE 2020 Operacional Programme for Competitiveness and Internationalisation (POCI), Portugal 2020, and Portuguese funds through Fundação para a Ciência e a Tecnologia (FCT)/Minist´erio da Ciência, Tecnologia e Ensino Superior in the framework of the project POCI-01-0145-FEDER-029471 (PTDC/BIA-BID/29471/2017) to T.J. Dantas. A.X. Carvalho, R. Gassmann, C.M.C. Abreu, and T.J. Dantas were supported by the FCT: CEECIND/01967/2017, CEECIND/00333/2017, CEECIND/01985/2018, and CEECIND/00771/2017, respectively. N. Vieira is a Junior Researcher under the scope of the FCT Transitional Rule DL57/2016, and her work was supported by a 2016 NARSAD Young Investigator Grant (#24929) from the Brain and Behavior Research Foundation. D.R.M. Rodrigues received a PhD fellowship from FCT (SFRH/BD/143985/2019) and support fromthe Biomedical Sciences PhD program at Instituto de Ciências Biom´edicas Abel Salazar (ICBAS). Some strains were provided by the National Bioresource Project for C. elegans and by the Caenorhabditis Genetics Center (CGC), which is funded by the National Institutes of Health Office of Research Infrastructure Programs (P40 OD010440).

Cilia are microtubule-based organelles that carry out a wide range of critical functions throughout the development of higher animals. Regardless of their type, all cilia rely on a motor-driven, bidirectional transport system known as intraflagellar transport (IFT). Of the many components of the IFT machinery, IFT20 is one of the smallest subunits. Nevertheless, IFT20 has been shown to play critical roles in the assembly of several types of mammalian cilia. Here we show that the IFT20 homolog in

Particulate fillers are often used to enhance the properties of soft materials; polymer composites often contain nanometer-sized particles to improve reinforcement, for example. The rationale for using nanometer-sized particles remains unclear, however, and classical micromechanical models cannot account for a scale dependent reinforcement. The systems studied here reveal that the reinforcement increases with decreasing filler size. A new relation is proposed, based only on the particle size, volume fraction, and relative moduli of filler and matrix, that describes the experimental results for reinforcement of both filled rubbers and a model system. (C) 2015 Elsevier Ltd. All rights reserved.

Polymer composite materials are widely used for their exceptional mechanical properties, notably their ability to resist large deformations. Here, we examine the failure stress and strain of rubbers reinforced by varying amounts of nano-sized silica particles. We find that small amounts of silica increase the fracture stress and strain, but too much filler makes the material become brittle and consequently fracture happens at small deformations. We thus find that as a function of the amount of filler there is an optimum in the breaking resistance at intermediate filler concentrations. We use a modified Griffith theory to establish a direct relation between the material properties and the fracture behavior that agrees with the experiment.

Context Glucose-insulin-potassium (GIK) infusion is a widely applicable, low-cost therapy that has been postulated to improve mortality in patients with acute ST-segment elevation myocardial infarction (STEMI). Given the potential global importance of GIK infusion, a large, adequately powered randomized trial is required to determine the effect of GIK on mortality in patients with STEMI. Objective To determine the effect of high-dose GIK infusion on mortality in patients with STEMI. Design, setting, and participants Randomized controlled trial conducted in 470 centers worldwide among 20,201 patients with STEMI who presented within 12 hours of symptom onset. The mean age of patients was 58.6 years, and evidence-based therapies were commonly used. Intervention Patients were randomly assigned to receive GIK intravenous infusion for 24 hours plus usual care (n = 10,091) or to receive usual care alone (controls; n = 10,110). Main outcome measures Mortality, cardiac arrest, cardiogenic shock, and reinfarction at 30 days after randomization. Results At 30 days, 976 control patients (9.7%) and 1004 GIK infusion patients (10.0%) died (hazard ratio [HR], 1.03; 95% confidence interval [CI], 0.95-1.13; P = .45). There were no significant differences in the rates of cardiac arrest (1.5% [151/10 107] in control and 1.4% [139/10,088] in GIK infusion; HR, 0.93; 95% CI, 0.74-1.17; P = .51), cardiogenic shock (6.3% [640/10 107] vs 6.6% [667/10 088]; HR, 1.05; 95% CI, 0.94-1.17; P = .38), or reinfarction (2.4% [246/10,107] vs 2.3% [236/10,088]; HR, 0.98; 95% CI, 0.82-1.17; P = .81). The rates of heart failure at 7 days after randomization were also similar between the groups (16.9% [1711/10,107] vs 17.1% [1721/10,088]; HR, 1.01; 95% CI, 0.95-1.08; P = .72). The lack of benefit of GIK infusion on mortality was consistent in prespecified subgroups, including in those with and without diabetes, in those presenting with and without heart failure, in those presenting early and later after symptom onset, and in those receiving and not receiving reperfusion therapy (thrombolysis or primary percutaneous coronary intervention). Conclusion In this large, international randomized trial, high-dose GIK infusion had a neutral effect on mortality, cardiac arrest, and cardiogenic shock in patients with acute STEMI.