O-glycosylation of the transcription factor SPATULA promotes style development in Arabidopsis.

O-linked ß-N-acetylglucosamine (O-GlcNAc) and O-fucose are two sugar-based post-translational modifications whose mechanistic role in plant signalling and transcriptional regulation is still largely unknown. Here we investigated how two O-glycosyltransferase enzymes of Arabidopsis thaliana, SPINDLY (SPY) and SECRET AGENT (SEC), promote the activity of the basic helix-loop-helix transcription factor SPATULA (SPT) during morphogenesis of the plant female reproductive organ apex, the style. SPY and SEC modify amino-terminal residues of SPT in vivo and in vitro by attaching O-fucose and O-GlcNAc, respectively. This post-translational regulation does not impact SPT homo- and heterodimerization events, although it enhances the affinity of SPT for the kinase PINOID gene locus and its transcriptional repression. Our findings offer a mechanistic example of the effect of O-GlcNAc and O-fucose on the activity of a plant transcription factor and reveal previously unrecognized roles for SEC and SPY in orchestrating style elongation and shape.

Differential sensitivity of ADF isovariants to a pH gradient promotes pollen tube growth.

The actin cytoskeleton is one of the targets of the pH gradient in tip-growing cells, but how cytosolic pH regulates the actin cytoskeleton remains largely unknown. We here demonstrate that Arabidopsis ADF7 and ADF10 function optimally at different pH levels when disassembling actin filaments. This differential pH sensitivity allows ADF7 and ADF10 to respond to the cytosolic pH gradient to regulate actin dynamics in pollen tubes. ADF7 is an unusual actin-depolymerizing factor with a low optimum pH in in vitro actin depolymerization assays. ADF7 plays a dominant role in promoting actin turnover at the pollen tube apex. ADF10 has a typically high optimum pH in in vitro assays and plays a dominant role in regulating the turnover and organization of subapical actin filaments. Thus, functional specification and cooperation of ADF isovariants with different pH sensitivities enable the coordination of the actin cytoskeleton with the cytosolic pH gradient to support pollen tube growth.

Activation of actin-depolymerizing factor by CDPK16-mediated phosphorylation promotes actin turnover in Arabidopsis pollen tubes.

As the stimulus-responsive mediator of actin dynamics, actin-depolymerizing factor (ADF)/cofilin is subject to tight regulation. It is well known that kinase-mediated phosphorylation inactivates ADF/cofilin. Here, however, we found that the activity of Arabidopsis ADF7 is enhanced by CDPK16-mediated phosphorylation. We found that CDPK16 interacts with ADF7 both in vitro and in vivo, and it enhances ADF7-mediated actin depolymerization and severing in vitro in a calcium-dependent manner. Accordingly, the rate of actin turnover is reduced in cdpk16 pollen and the amount of actin filaments increases significantly at the tip of cdpk16 pollen tubes. CDPK16 phosphorylates ADF7 at Serine128 both in vitro and in vivo, and the phospho-mimetic mutant ADF7S128D has enhanced actin-depolymerizing activity compared to ADF7. Strikingly, we found that failure in the phosphorylation of ADF7 at Ser128 impairs its function in promoting actin turnover in vivo, which suggests that this phospho-regulation mechanism is biologically significant. Thus, we reveal that CDPK16-mediated phosphorylation up-regulates ADF7 to promote actin turnover in pollen.

Carbon Dots-Based Fluorescence Assay for the Facile and Reliable Detection of Ag+ in Natural Water and Serum Samples.

In this report, red-emissive carbon dots (C-dots) were facilely prepared from o-phenylenediamine via microwave-assisted hydrothermal treatment. The C-dots demonstrated excitation wavelength-independent emission with maximums at 621 nm that could be effectively quenched by Ag+ via static quenching. This phenomenon was exploited to establish a sensitive fluorescence assay with a low detection limit (0.37 µM) and wide linear range (0-50 µM). In addition, this assay demonstrated excellent selectivity toward Ag+, free from the interference of 16 commonly seen metal ions. Most importantly, the assay demonstrated high reliability toward samples in deionized water, mineral water, lake water, and serum, which could indicate potential applications for Ag+ monitoring in complicated natural and biological environments.

Indocyanine green derived carbon dots with significantly enhanced properties for efficient photothermal therapy.

A simple yet effective strategy to enhance the properties of traditional dye indocyanine green (ICG) in all aspects was proposed and demonstrated. Specifically, indocyanine green-derived carbon dots (ICGCDs) were synthesized from ICG via a simple hydrothermal treatment. The ICGCDs exhibited significantly enhanced thermal stability and anti-photobleaching compared to ICG. Furthermore, their photothermal properties were also notably strengthened, in which a wider functional pH range, 50% improvement in photothermal conversion efficiency and superior photothermal cyclability were achieved. Thanks to these superior properties, ICGCDs were demonstrated as efficient NIR bioimaging and photothermal agents in both in vitro and in vivo experiments. Most excitingly, the strategy demonstrated in this study is likely to have broad applications in other systems.

Calcium signaling in plant immunity: a spatiotemporally controlled symphony.

Calcium ions (Ca2+) are prominent intracellular messengers in all eukaryotic cells. Recent studies have emphasized the crucial roles of Ca2+ in plant immunity. Here, we review the latest progress on the spatiotemporal control of Ca2+ function in plant immunity. We discuss discoveries of how Ca2+ influx is triggered upon the activation of immune receptors, how Ca2+-permeable channels are activated, how Ca2+ signals are decoded inside plant cells, and how these signals are switched off. Despite recent advances, many open questions remain and we highlight the existing toolkit and the new technologies to address the outstanding questions of Ca2+ signaling in plant immunity.

Hierarchical framework for mobile robots to effectively and autonomously explore unknown environments.

Achieving efficient and safe autonomous exploration in unknown environments is an urgent challenge to be overcome in the field of robotics. Existing exploration methods based on random and greedy strategies cannot ensure that the robot moves to the unknown area as much as possible, and the exploration efficiency is not high. In addition, because the robot is located in an unknown environment, the robot cannot obtain enough information to process the surrounding environment and cannot guarantee absolute safety. To improve the efficiency and safety of exploring unknown environments, we propose an autonomous exploration motion planning framework that is divided into the exploration and obstacle avoidance levels. The two levels are independent and interconnected. The exploration level finds the optimal frontier target point in the global scope based on the forward filtering angle and cost function, attracting the robot to move to the unknown area as much as possible, and improving the exploration efficiency; the obstacle avoidance level establishes a scenario-speed conversion mechanism, and the target point and obstacle information are weighed to realise dynamic motion planning and completes obstacle avoidance control, and ensures the safety of exploration. Experiments in different simulation scenarios and real environments verify the superiority of the method. Results show that our method is superior to the existing methods.

Sensitive, Selective and Reliable Detection of Fe3+ in Lake Water via Carbon Dots-Based Fluorescence Assay.

In this study, C-dots were facilely synthesized via microwave irradiation using citric acid and ethylenediamine as carbon precursors. The fluorescence emissions of the C-dots could be selectively quenched by Fe3+, and the degree of quenching was linearly related to the concentrations of Fe3+ presented. This phenomenon was utilized to develop a sensitive fluorescence assay for Fe3+ detection with broad linear range (0-250, 250-1200 µmol/L) and low detection limit (1.68 µmol/L). Most importantly, the assay demonstrated high reliability towards samples in deionized water, tap water and lake water, which should find potential applications for Fe3+ monitoring in complicated environments.

Floral symmetry: the geometry of plant reproduction.

The flower is an astonishing innovation that arose during plant evolution allowing flowering plants - also known as angiosperms - to dominate life on earth in a relatively short period of geological time. Flowers are formed from secondary meristems by co-ordinated differentiation of flower organs, such as sepals, petals, stamens, and carpels. The position, number and morphology of these flower organs impose a geometrical pattern - or symmetry type - within the flower which is a trait tightly connected to successful reproduction. During evolution, flower symmetry switched from the ancestral poly-symmetric (radial symmetry) to the mono-symmetric (bilateral symmetry) type multiple times, including numerous reversals, with these events linked to co-evolution with pollinators and reproductive strategies. In this review, we introduce the diversity of flower symmetry, trace its evolution in angiosperms, and highlight the conserved genetic basis underpinning symmetry control in flowers. Finally, we discuss the importance of building upon the concept of flower symmetry by looking at the mechanisms orchestrating symmetry within individual flower organs and summarise the current scenario on symmetry patterning of the female reproductive organ, the gynoecium, the ultimate flower structure presiding over fertilisation and seed production.

Research and Optimization of Process Parameters for Internal Thread Forming Based on Numerical Simulation and Experimental Analysis.

In order to improve the forming quality of extruded thread, finite element analysis and experimental research are combined to reduce the two keys that affect thread quality in the machining process-extrusion torque and extrusion temperature. The effects of different processing parameters on the extrusion torque and temperature are obtained by numerical simulation, including the bottom hole diameter of the workpiece, the machine tool speed, and the lubrication medium. For the purpose of reducing extrusion torque and temperature, the process parameters for internal thread forming are further optimized by orthogonal design. It is determined that when machining the M22 × 2 internal thread on the connecting rod of the marine diesel engine made of 42CrMo4 steel, the bottom hole diameter of the workpiece should be 21.20 mm, the speed of the machine tool should be 40 RPM, and the lubricating medium should be PDMS polydimethylsiloxane coolant. Compared to before optimization, the maximum extrusion torque and the maximum extrusion temperature are reduced by 19.27% and 15.07%, respectively. On the premise of ensuring the thread connection strength, the height of the thread tooth is reduced by 0.052 mm, and the surface condition of the thread is improved. The surface microhardness at the root, top, and side of the thread increases by about 5 HV0.2, and the depth of the hardened layer increases by 0.05 mm. The results show that the quality of the optimized thread is higher.

Angiogenin and plexin-B2 axis promotes glioblastoma progression by enhancing invasion, vascular association, proliferation and survival.

BACKGROUND: Angiogenin is a multifunctional secreted ribonuclease that is upregulated in human cancers and downregulated or mutationally inactivated in neurodegenerative diseases. A role for angiogenin in glioblastoma was inferred from the inverse correlation of angiogenin expression with patient survival but had not been experimentally investigated. METHODS: Angiogenin knockout mice were generated and the effect of angiogenin deficiency on glioblastoma progression was examined. Angiogenin and plexin-B2 genes were knocked down in glioblastoma cells and the changes in cell proliferation, invasion and vascular association were examined. Monoclonal antibodies of angiogenin and small molecules were used to assess the therapeutic activity of the angiogenin-plexin-B2 pathway in both genetic and xenograft animal models. RESULTS: Deletion of Ang1 gene prolonged survival of PDGF-induced glioblastoma in mice in the Ink4a/Arf-/-:Pten-/- background, accompanied by decreased invasion, vascular association and proliferation. Angiogenin upregulated MMP9 and CD24 leading to enhanced invasion and vascular association. Inhibition of angiogenin or plexin-B2, either by shRNA, monoclonal antibody or small molecule inhibitor, decreases sphere formation of patient-derived glioma stem cells, reduces glioblastoma proliferation and invasion and inhibits glioblastoma growth in both genetic and xenograft animal models. CONCLUSIONS: Angiogenin and its receptor, plexin-B2, are a pair of novel regulators that mediate invasion, vascular association and proliferation of glioblastoma cells. Inhibitors of the angiogenin-plexin-B2 axis have therapeutic potential against glioblastoma.

Wnt signaling polarizes cortical actin polymerization to increase daughter cell asymmetry.

Asymmetric positioning of the mitotic spindle contributes to the generation of two daughter cells with distinct sizes and fates. Here, we investigated an asymmetric division in the Caenorhabditis elegans Q neuroblast lineage. In this division, beginning with an asymmetrically positioned spindle, the daughter-cell size differences continuously increased during cytokinesis, and the smaller daughter cell in the posterior eventually underwent apoptosis. We found that Arp2/3-dependent F-actin assembled in the anterior but not posterior cortex during division, suggesting that asymmetric expansion forces generated by actin polymerization may enlarge the anterior daughter cell. Consistent with this, inhibition of cortical actin polymerization or artificially equalizing actin assembly led to symmetric cell division. Furthermore, disruption of the Wnt gradient or its downstream components impaired asymmetric cortical actin assembly and caused symmetric division. Our results show that Wnt signaling establishes daughter cell asymmetry by polarizing cortical actin polymerization in a dividing cell.

Functional non-equivalence of pollen ADF isovariants in Arabidopsis.

ADF/cofilin is a central regulator of actin dynamics. We previously demonstrated that two closely related Arabidopsis class IIa ADF isovariants, ADF7 and ADF10, are involved in the enhancement of actin turnover in pollen, but whether they have distinct functions remains unknown. Here, we further demonstrate that they exhibit distinct functions in regulating actin turnover both in vitro and in vivo. We found that ADF7 binds to ADP-G-actin with lower affinity, and severs and depolymerizes actin filaments less efficiently in vitro than ADF10. Accordingly, in pollen grains, ADF7 more extensively decorates actin filaments and is less freely distributed in the cytoplasm compared to ADF10. We further demonstrate that ADF7 and ADF10 show distinct intracellular localizations during pollen germination, and they have non-equivalent functions in promoting actin turnover in pollen. We thus propose that cooperation and labor division of ADF7 and ADF10 enable pollen cells to achieve exquisite control of the turnover of different actin structures to meet different cellular needs.

Metabolite Characteristics Analysis of Siliques and Effects of Lights on the Accumulation of Glucosinolates in Siliques of Rapeseed.

Glucosinolates (GSLs) are naturally occurring secondary metabolites found in the Brassicaceae family, which mainly synthesize in the siliques with a wide range of functions. In this study, we investigated the effects of lights on metabolites in siliques of rapeseed through ultra high-performance liquid chromatography (UPLC)-heated electrospray ionization (HESI)-tandem mass spectrometry (MS/MS). A total of 249 metabolites, including 29 phenolic acids, 38 flavonoids, 22 GSLs, 93 uncalculated and 67 unknown compounds, were identified in siliques of rapeseed. Meanwhile, 62 metabolites showed significant differences after shading treatment, which were mainly GSLs and unknown compounds. Interestingly, the amounts of 10 GSLs had high accumulation levels in siliques, while the expression levels of their corresponding biosynthetic genes (AOP, GSL-OH, IGMT, and ST5a) were obviously reduced after shading treatment. Further evidence showed that the amounts of GSLs were significantly reduced in seeds, in accordance with the expression profiles of transporter genes (BnaGTRs). Our findings indicated that lights could affect the accumulation and transportation of GSLs from siliques to seeds in rapeseed. Therefore, this study facilitates a better understanding of metabolic characteristics of siliques and provides insight into the importance of light for GSLs accumulation and transportation in siliques and seeds of rapeseed.

Prioritizing de novo autism risk variants with calibrated gene- and variant-scoring models.

Whole-exome and whole-genome sequencing studies in autism spectrum disorder (ASD) have identified hundreds of thousands of exonic variants. Only a handful of them, primarily loss-of-function variants, have been shown to increase the risk for ASD, while the contributory roles of other variants, including most missense variants, remain unknown. New approaches that combine tissue-specific molecular profiles with patients' genetic data can thus play an important role in elucidating the functional impact of exonic variation and improve understanding of ASD pathogenesis. Here, we integrate spatio-temporal gene co-expression networks from the developing human brain and protein-protein interaction networks to first reach accurate prioritization of ASD risk genes based on their connectivity patterns with previously known high-confidence ASD risk genes. We subsequently integrate these gene scores with variant pathogenicity predictions to further prioritize individual exonic variants based on the positive-unlabeled learning framework with gene- and variant-score calibration. We demonstrate that this approach discriminates among variants between cases and controls at the high end of the prediction range. Finally, we experimentally validate our top-scoring de novo mutation NP_001243143.1:p.Phe309Ser in the sodium/potassium-transporting ATPase ATP1A3 to disrupt protein binding with different partners.

Microstructure and Wear Resistance of Laser-Clad Ni-Cu-Mo-W-Si Coatings on a Cu-Cr-Zr Alloy.

To improve the wear resistance of high-strength and high-conductivity Cu-Cr-Zr alloys in high-speed and heavy load friction environments, coatings including Ni-Cu, Ni-Cu-10(W,Si), Ni-Cu-10(Mo,W,Si), and Ni-Cu-15(Mo,W,Si) (with an atomic ratio of Mo,W to Si of 1:2) were prepared using coaxial powder-feeding laser cladding technology. The microstructure and wear performance of coatings were chiefly investigated. The results revealed that (Mo,W)Si2 and MoNiSi phases are found in the Ni-Cu-10(Mo,W,Si) and Ni-Cu-15(Mo,W,Si) coating. WSi2 phases are found in the Ni-Cu-10(W,Si) coating. The degree of grain refinement in Ni-Cu-10(Mo,W,Si) was greater than that of the Ni-Cu-10(W,Si) coating after the effect of Mo. The excellent wear resistance and micro-hardness of the Ni-Cu-15(Mo,W,Si) coating were attributed to the increase in its dispersion phase, which were approximately 34.72 mg/km and 428 HV, 27.1% and 590% higher than the Cu-Cr-Zr substrate, respectively. The existence of silicide plays an important role in grain refinement due to the promotion of nucleation and the inhibition of grain growth. In addition, the wear mechanism transformed from adhesive wear in the Ni-Cu coating with no silicides to abrasive wear in the Ni-Cu-15(Mo,W,Si) coating with high levels of silicides.

Actin filament debranching regulates cell polarity during cell migration and asymmetric cell division.

The formation of the branched actin networks is essential for cell polarity, but it remains unclear how the debranching activity of actin filaments contributes to this process. Here, we showed that an evolutionarily conserved coronin family protein, the Caenorhabditis elegans POD-1, debranched the Arp2/3-nucleated actin filaments in vitro. By fluorescence live imaging analysis of the endogenous POD-1 protein, we found that POD-1 colocalized with Arp2/3 at the leading edge of the migrating C. elegans neuroblasts. Conditional mutations of POD-1 in neuroblasts caused aberrant actin assembly, disrupted cell polarity, and impaired cell migration. In C. elegans one-cell-stage embryos, POD-1 and Arp2/3, moved together during cell polarity establishment, and inhibition of POD-1 blocked Arp2/3 motility and affected the polarized cortical flow, leading to symmetric segregation of cell fate determinants. Together, these results indicate that F-actin debranching organizes actin network and cell polarity in migrating neuroblasts and asymmetrically dividing embryos.

The CAFA challenge reports improved protein function prediction and new functional annotations for hundreds of genes through experimental screens.

BACKGROUND: The Critical Assessment of Functional Annotation (CAFA) is an ongoing, global, community-driven effort to evaluate and improve the computational annotation of protein function. RESULTS: Here, we report on the results of the third CAFA challenge, CAFA3, that featured an expanded analysis over the previous CAFA rounds, both in terms of volume of data analyzed and the types of analysis performed. In a novel and major new development, computational predictions and assessment goals drove some of the experimental assays, resulting in new functional annotations for more than 1000 genes. Specifically, we performed experimental whole-genome mutation screening in Candida albicans and Pseudomonas aureginosa genomes, which provided us with genome-wide experimental data for genes associated with biofilm formation and motility. We further performed targeted assays on selected genes in Drosophila melanogaster, which we suspected of being involved in long-term memory. CONCLUSION: We conclude that while predictions of the molecular function and biological process annotations have slightly improved over time, those of the cellular component have not. Term-centric prediction of experimental annotations remains equally challenging; although the performance of the top methods is significantly better than the expectations set by baseline methods in C. albicans and D. melanogaster, it leaves considerable room and need for improvement. Finally, we report that the CAFA community now involves a broad range of participants with expertise in bioinformatics, biological experimentation, biocuration, and bio-ontologies, working together to improve functional annotation, computational function prediction, and our ability to manage big data in the era of large experimental screens.

AP3M harbors actin filament binding activity that is crucial for vacuole morphology and stomatal closure in Arabidopsis.

Stomatal movement is essential for plant growth. This process is precisely regulated by various cellular activities in guard cells. F-actin dynamics and vacuole morphology are both involved in stomatal movement. The sorting of cargoes by clathrin adaptor protein (AP) complexes from the Golgi to the vacuole is critical for establishing a normal vacuole morphology. In this study, we demonstrate that the medium subunit of the AP3 complex (AP3M) binds to and severs actin filaments in vitro and that it participates in the sorting of cargoes (such as the sucrose exporter SUC4) to the tonoplast, and thereby regulates stomatal closure in Arabidopsis thaliana Defects in AP3 or SUC4 led to more rapid water loss and delayed stomatal closure, as well as hypersensitivity to drought stress. In ap3m mutants, the F-actin status was altered compared to the wild type, and the sorted cargoes failed to localize to the tonoplast. AP3M contains a previously unidentified F-actin binding domain that is conserved in AP3M homologs in both plants and animals. Mutations in the F-actin binding domain of AP3M abolished its F-actin binding activity in vitro, leading to an aberrant vacuole morphology and reduced levels of SUC4 on the tonoplast in guard cells. Our findings indicate that the F-actin binding activity of AP3M is required for the precise localization of AP3-dependent cargoes to the tonoplast and for the regulation of vacuole morphology in guard cells during stomatal closure.

CAGI SickKids challenges: Assessment of phenotype and variant predictions derived from clinical and genomic data of children with undiagnosed diseases.

Whole-genome sequencing (WGS) holds great potential as a diagnostic test. However, the majority of patients currently undergoing WGS lack a molecular diagnosis, largely due to the vast number of undiscovered disease genes and our inability to assess the pathogenicity of most genomic variants. The CAGI SickKids challenges attempted to address this knowledge gap by assessing state-of-the-art methods for clinical phenotype prediction from genomes. CAGI4 and CAGI5 participants were provided with WGS data and clinical descriptions of 25 and 24 undiagnosed patients from the SickKids Genome Clinic Project, respectively. Predictors were asked to identify primary and secondary causal variants. In addition, for CAGI5, groups had to match each genome to one of three disorder categories (neurologic, ophthalmologic, and connective), and separately to each patient. The performance of matching genomes to categories was no better than random but two groups performed significantly better than chance in matching genomes to patients. Two of the ten variants proposed by two groups in CAGI4 were deemed to be diagnostic, and several proposed pathogenic variants in CAGI5 are good candidates for phenotype expansion. We discuss implications for improving in silico assessment of genomic variants and identifying new disease genes.