Full Assembly Research Articles

Phosphatidylinositol and phosphatidylinositol-3-phosphate activate HOPS to catalyze SNARE assembly, allowing small headgroup lipids to support the terminal steps of membrane fusion.

Intracellular membrane fusion requires Rab GTPases, tethers, SNAREs of the R, Qa, Qb, and Qc families, and SNARE chaperones of the Sec17 (SNAP), Sec18 (NSF), and SM (Sec1/Munc18) families. The vacuolar HOPS complex combines the functions of membrane tethering and SM catalysis of SNARE assembly. HOPS is activated for this catalysis by binding to the vacuolar lipids and Rab. Of the eight major vacuolar lipids, we now report that phosphatidylinositol and phosphatidylinositol-3-phosphate are required to activate HOPS for SNARE complex assembly. These lipids plus ergosterol also allow full trans-SNARE complex assembly, yet do not support fusion, which is reliant on either phosphatidylethanolamine (PE) or on phosphatidic acid (PA), phosphatidylserine (PS), and diacylglycerol (DAG). Fusion with a synthetic tether and without HOPS, or even without SNAREs, still relies on either PE or on PS, PA, and DAG. These lipids are thus required for the terminal bilayer rearrangement step of fusion, distinct from the lipid requirements for the earlier step of activating HOPS for trans-SNARE assembly.

Method for Rapid Modeling of Distortion in Laser Powder Bed Fusion Metal Additive Manufacturing Parts

The simulation and modeling of part-level distortion and residual stress in diverse metal additive manufacturing (AM) geometries has great potential to enable the rapid adoption of this technology in engineering design. Moreover, the use of additive manufacturing component libraries (CLs) offer a computationally efficient means of quantifying these part-level defects resultant from AM processing. We report on how the individual simulations of simple shapes, potential entries in a CL, can be superimposed to provide an indication of distortion and residual stresses in complex geometries. Laser powder bed fusion AM was used to construct test geometries of varied shapes and their combinations in the form of CLs in an effort to characterize location-dependent and feature-dependent distortion distributions. Blue light scanning was used to experimentally measure 3D distortions in order to investigate the interaction between the component shapes and local boundary conditions. Overall, part-level distortions were highly dependent on test component geometry, local boundary conditions, and shape combination. Commercial finite element software was used to verify experimental trends and to make predictions of distortion. The use of CLs resulted in over 20 times savings in computational cost while reproducing overall trends in distortion for test geometry assemblies. Therefore, it is anticipated that the use of CLs for L-PBF AM geometries has demonstrated potential to facilitate efficient simulations of full component AM assemblies, thereby reducing the need for costly trial-and-error-type experimental analysis.

Comparison of design approaches for low-cost sampling mechanisms in open-source chemical instrumentation

Robotic positioning systems are used in a variety of chemical instruments, primarily for liquid handling purposes, such as autosamplers from vials or well plates. Here, two approaches to the design of open-source autosampler positioning systems for use with 96-well plates are described and compared. The first system, a 3-axis design similar to many low-cost 3D printers that are available on the market, is constructed using an aluminum frame and stepper motors. The other system relies upon a series of 3D printed parts to achieve movement with a series of linker arms based on Selective Compliance Assembly Robot Arm (SCARA) design principles. Full printer design files, assembly instructions, software, and user directions are included for both samplers. The positioning precision of the 3-axis system is better than the SCARA mechanism due to finer motor control, albeit with a slightly higher cost of materials. Based on the improved precision of this approach, the 3-axis autosampler system was used to demonstrate the generation of a segmented flow droplet stream from adjacent wells within a 96-well plate.

Heat Transfer Performance of Cu-Cr-Zr Tube with Swirl Insert Under Cyclic Thermal Loading in Monoblock Divertor

Development and optimization of the plasma-facing components for the fusion reactors ITER and DEMO are necessary for sufficient heat removal because of the high heat fluxes in these systems. In this work, we consider the heat transfer performance of the Cu-Cr-Zr alloy tube with a swirl (twisted tape) insert within a monoblock divertor experiencing cyclic thermal loading expected during ITER operating conditions. Thermal loading is examined up to 2000 cycles, leading to increased tube surface roughness and decreased tube thermal conductivity. A simplified model of thermal-hydraulic performance is used that accounts for forced convection in the swirled flow, conduction through the Cu-Cr-Zr tube, and tube fouling (surface roughness and thermal conductivity changes). From our work, it is found that the overall heat transfer rate of the tube is enhanced with increased thermal loading over a wide range of Reynolds numbers (i.e., flow rates). This is due to the increase of convective heat transfer from turbulence enhancement induced by increasing surface roughness. However, the increase in surface roughness also leads to an increase in pressure losses in the system, requiring increased pumping power to maintain flow rates. We consider the heat transfer rate at equivalent pumping power (quantified by the overall enhancement ratio) and find it has a complicated dependence on Reynolds number and the number of thermal loading cycles. In particular, we see that for a Reynolds number of 1 000 000, the overall enhancement ratio is decreased by up to 9% at 2000 loading cycles. Such a decrease could meaningfully impact the operations of ITER or DEMO, requiring additional pumping input to maintain sufficient heat removal. This suggests the need for further investigation of the thermal-hydraulic performance of plasma-facing components, including the full monoblock assembly, after many loading cycles.

Design and Optimization of Rear Wheel Assembly for All-Terrain Vehicle

This paper describes the design and optimisation of rear wheel assembly for an all-terrain vehicle. The components of rear wheel assembly are designed with an optimised design intent with light weight material selection. The designed assembly is then discretised using finite element modelling through Hexa-dominant 3D mesh. The upright knuckle and the hub are assessed for their integrity check and the performance of full assembly is verified through simulation. From the post-processed stress results, factor of safety for the designed parts is presented. The presented work can be expanded to optimise the upright and the hub based the resolved gradient stress plots from the finite element analysis.

Assembly of γ-secretase occurs through stable dimers after exit from the endoplasmic reticulum.

γ-Secretase affects many physiological processes through targeting >100 substrates; malfunctioning links γ-secretase to cancer and Alzheimer's disease. The spatiotemporal regulation of its stoichiometric assembly remains unresolved. Fractionation, biochemical assays, and imaging support prior formation of stable dimers in the ER, which, after ER exit, assemble into full complexes. In vitro ER budding shows that none of the subunits is required for the exit of others. However, knockout of any subunit leads to the accumulation of incomplete subcomplexes in COPII vesicles. Mutating a DPE motif in presenilin 1 (PSEN1) abrogates ER exit of PSEN1 and PEN-2 but not nicastrin. We explain this by the preferential sorting of PSEN1 and nicastrin through Sec24A and Sec24C/D, respectively, arguing against full assembly before ER exit. Thus, dimeric subcomplexes aided by Sec24 paralog selectivity support a stepwise assembly of γ-secretase, controlling final levels in post-Golgi compartments.

Nanopore-based DNA sequencing sensors and CMOS readout approaches

PurposeNanopore-based molecular sensing and measurement, specifically DNA sequencing, is advancing at a fast pace. Some embodiments have matured from coarse particle counters to enabling full human genome assembly. This evolution has been powered not only by improvements in the sensors themselves, but also in the assisting microelectronic CMOS readout circuitry closely interfaced to them. In this light, this paper aims to review established and emerging nanopore-based sensing modalities considered for DNA sequencing and CMOS microelectronic methods currently being used.Design/methodology/approachReadout and amplifier circuits, which are potentially appropriate for conditioning and conversion of nanopore signals for downstream processing, are studied. Furthermore, arrayed CMOS readout implementations are focused on and the relevant status of the nanopore sensor technology is reviewed as well.FindingsIon channel nanopore devices have unique properties compared with other electrochemical cells. Currently biological nanopores are the only variants reported which can be used for actual DNA sequencing. The translocation rate of DNA through such pores, the current range at which these cells operate on and the cell capacitance effect, all impose the necessity of using low-noise circuits in the process of signal detection. The requirement of using in-pixel low-noise circuits in turn tends to impose challenges in the implementation of large size arrays.Originality/valueThe study presents an overview on the readout circuits used for signal acquisition in electrochemical cell arrays and investigates the specific requirements necessary for implementation of nanopore-type electrochemical cell amplifiers and their associated readout electronics.

Design and analysis of a Cross Car frame. Proposal of a three-level appraisal methodology

This paper analyses the structural design process of a Cross Car frame. Existing international and national technical regulations were analysed. The analysis of the regulatory guidelines led to the conclusion that the design only includes geometric or material restrictions at the international level. There are other more demanding national regulations (Spanish, for instance) that include structural verifications through static calculations for unidirectional actions (vertical, longitudinal or lateral). The main contribution of this paper is a novel three-level appraisal and a proposed redesign methodology. At the first level, a geometric and material verification is carried out. The second level involves the verification under unidirectional static actions. The third level entails a dynamic verification of three-dimensional combined actions. The load case is obtained from the computer multibody dynamics simulation of the full vehicle assembly in the worst case of driving conditions on circuits. This methodology is a far more detailed tool than traditional design processes. The use of this methodology allows for design optimisation, including all the effects of powertrain, brakes, suspension, steering and driver.

Волонтёрская деятельность как фактор формирования профессиональных компетенций (на примере лекторской группы Рязанского филиала Московского университета МВД России имени В. Я. Кикотя)

This article is devoted to the description of the experience of the volunteer activities of lecturer groups of the Ryazan branch of the V. Kikot Moscow University of the Ministry of the Interior of Russia. Peculiarities of forming professional psychological and pedagogical competencies of law students and cadets of universities the Ministry of the Interior, while giving public lectures, are illustrated. Cadets study in details the subject on which they are going to speak to schoolchildren, study their psychological and age characteristics, psychology of public speech, the basics of eloquence. Future lecturers do some kind of test run in front of other cadets, after that they have the possibility to analyse their own mistakes righting wrongs. While real public lectures, cadets study to speak in front of various categories of audience: in front of a group of several people, one school grade or in a full assembly hall. Lectures on the same topic can be held for students of different ages and for different categories of children: for students of ordinary schools, boarding schools, gymnasiums and lyceums; for law-abiding children and for those who are registered by juvenile departments. The article shows how giving public lectures in a lecturer group can form the professional competences of a law enforcement officer.

Comprehensive identification of transposable element insertions using multiple sequencing technologies

Transposable elements (TEs) help shape the structure and function of the human genome. When inserted into some locations, TEs may disrupt gene regulation and cause diseases. Here, we present xTea (x-Transposable element analyzer), a tool for identifying TE insertions in whole-genome sequencing data. Whereas existing methods are mostly designed for short-read data, xTea can be applied to both short-read and long-read data. Our analysis shows that xTea outperforms other short read-based methods for both germline and somatic TE insertion discovery. With long-read data, we created a catalogue of polymorphic insertions with full assembly and annotation of insertional sequences for various types of retroelements, including pseudogenes and endogenous retroviruses. Notably, we find that individual genomes have an average of nine groups of full-length L1s in centromeres, suggesting that centromeres and other highly repetitive regions such as telomeres are a significant yet unexplored source of active L1s. xTea is available at https://github.com/parklab/xTea.

Full assembly of HIV-1 particles requires assistance of the membrane curvature factor IRSp53.

During HIV-1 particle formation, the requisite plasma membrane curvature is thought to be solely driven by the retroviral Gag protein. Here, we reveal that the cellular I-BAR protein IRSp53 is required for the progression of HIV-1 membrane curvature to complete particle assembly. siRNA-mediated knockdown of IRSp53 gene expression induces a decrease in viral particle production and a viral bud arrest at half completion. Single-molecule localization microscopy at the cell plasma membrane shows a preferential localization of IRSp53 around HIV-1 Gag assembly sites. In addition, we observe the presence of IRSp53 in purified HIV-1 particles. Finally, HIV-1 Gag protein preferentially localizes to curved membranes induced by IRSp53 I-BAR domain on giant unilamellar vesicles. Overall, our data reveal a strong interplay between IRSp53 I-BAR and Gag at membranes during virus assembly. This highlights IRSp53 as a crucial host factor in HIV-1 membrane curvature and its requirement for full HIV-1 particle assembly.

Research on Optimizing Selection and Optimizing Matching Technologies of Aeroengine Fan Rotor Blades

Aiming at the problem of low resources utilization of rotating blades in the selection process of aeroengine fan rotor blades, this paper takes the first-order bending dispersion, first-order torque dispersion, and gravitational moment difference of rotor blades as the selection criteria and takes the minimum remaining blades as the optimization goal. An intelligent selection algorithm of blades based on the collocation degree of blades is proposed and achieves the efficient selection and full utilization of rotating blades. Aiming at the problem of multiple installations and multiple adjustments and low assembly success rate of fan rotor blades, this paper takes the gravity moment difference of the two blades at the diagonal position of 180° as the constraint and takes the minimum residual unbalance as the optimization objective, adopts the improved simulated annealing algorithm to optimize the assembly sequence of rotating blades, and greatly reduces the residual unbalance of blades, which is beneficial to reduce the number of assembly adjustments of blades. The optimizing selection and optimizing matching methods of rotating blades realize the full utilization and efficient assembly of blades and lays a foundation for the reliability and robustness of the assembly quality and service performance of blades.

Flow Physics During Surge and Recovery of a Multi-Stage High-Speed Compressor

Abstract Stall followed by surge in a high-speed compressor can lead to violent disruption of the flow, damage to the blade structures and eventually engine shutdown. Knowledge of unsteady blade loading during surge is crucial for compressor design such as axial gap optimization. The aim of this paper is to demonstrate the feasibility of using three-dimensional full assembly unsteady Reynolds-averaged Navier–Stokes (URANS) CFD for modeling surge cycles of an eight-stage high-speed compressor rig. Results from this work show stalling of the mid-stages is the surge trigger. During the flow reversal, a strong acoustic reflection occurs when the convected entropy perturbations reach the intake opening, which increase the blade loading significantly. During recovery, a hysteresis loop was recorded due to hot air re-ingestion, which led to a strong shear at mid-span of the inlet guide vane (IGV)/R1 domain and the formation of rotating helical flow structures. The final phase of recovery was accompanied by a four-cell multi-row tip rotating stall, which was cleared as the compressor recovered to the forward flow characteristic. It was also shown that the single passage model, despite its limitations and shortcomings in modeling recovery, can predict reasonably accurate transient flow features during surge and thus provide considerable insight to the flow behavior, which can be used to obtain a first approximation of casing and blade loading.

Equivalent Linearization of Bladed Disk Assemblies With Friction Nonlinearities Under Random Excitation

AbstractThis article addresses the vibrational behavior of bladed disk assemblies with nonlinear shroud coupling under random excitation. In order to increase the service life and safety of turbine blades, intense calculations are carried out to predict the vibrational behavior. The use of friction dampers for energy dissipation and suppression of large amplitudes makes the mechanical system nonlinear, which complicates the calculations. Depending on the stage, different types of excitation can occur in a turbine, from clearly defined deterministic to random excitation. So far, the latter problem has only been dealt with to a limited extent in the literature on turbomachinery. Nevertheless, there are in general different approaches and methods to address this problem most of which are strongly restricted with regard to the number of degrees-of-freedom (DOF). The focus of this paper is the application of an equivalent linearization method (ELM) to calculate the stochastic response of an academic model of a bladed disk assembly under random excitation. The nonlinear contact is modeled both with an elastic Coulomb-slider and a Bouc–Wen formulation to reproduce the hysteretic character of a friction nonlinearity occurring in the presence of a friction damper. Both the excitation and the response are limited to mean-free, stationary stochastic processes, which means that the stochastic moments do not change over time. Unlike previous papers on this topic, the calculations are performed on a full bladed disk assembly in which each segment is approximated with several degrees-of-freedom.

Reconstruction of the Origin of a Neo-Y Sex Chromosome and Its Evolution in the Spotted Knifejaw, Oplegnathus punctatus.

Sex chromosomes are a peculiar constituent of the genome because the evolutionary forces that fix the primary sex-determining gene cause genic degeneration and accumulation of junk DNA in the heterogametic partner. One of the most spectacular phenomena in sex chromosome evolution is the occurrence of neo-Y chromosomes, which lead to X1X2Y sex-determining systems. Such neo-sex chromosomes are critical for understanding the processes of sex chromosome evolution because they rejuvenate their total gene content. We assembled the male and female genomes at the chromosome level of the spotted knifejaw (Oplegnathus punctatus), which has a cytogenetically recognized neo-Y chromosome. The full assembly and annotation of all three sex chromosomes allowed us to reconstruct their evolutionary history. Contrary to other neo-Y chromosomes, the fusion to X2 is quite ancient, estimated at 48 Ma. Despite its old age and being even older in the X1 homologous region which carries a huge inversion that occurred as early as 55–48 Ma, genetic degeneration of the neo-Y appears to be only moderate. Transcriptomic analysis showed that sex chromosomes harbor 87 genes, which may serve important functions in the testis. The accumulation of such male-beneficial genes, a large inversion on the X1 homologous region and fusion to X2 appear to be the main drivers of neo-Y evolution in the spotted knifejaw. The availability of high-quality assemblies of the neo-Y and both X chromosomes make this fish an ideal model for a better understanding of the variability of sex determination mechanisms and of sex chromosome evolution.

The Accessory Helix of Complexin Stabilizes a Partially Unzippered State of the SNARE Complex and Mediates the Complexin Clamping Function In Vivo.

Spontaneous synaptic transmission is regulated by the protein complexin (Cpx). Cpx binds the SNARE complex, a coil-coiled four-helical bundle that mediates the attachment of a synaptic vesicle (SV) to the presynaptic membrane (PM). Cpx is thought to clamp spontaneous fusion events by stabilizing a partially unraveled state of the SNARE bundle; however, the molecular detail of this mechanism is still debated. We combined electrophysiology, molecular modeling, and site-directed mutagenesis in Drosophila to develop and validate the atomic model of the Cpx-mediated clamped state of the SNARE complex. We took advantage of botulinum neurotoxins (BoNTs) B and G, which cleave the SNARE protein synaptobrevin (Syb) at different sites. Monitoring synaptic depression on BoNT loading revealed that the clamped state of the SNARE complex has two or three unraveled helical turns of Syb. Site-directed mutagenesis showed that the Cpx clamping function is predominantly maintained by its accessory helix (AH), while molecular modeling suggested that the Cpx AH interacts with the unraveled C terminus of Syb and the SV lipid bilayer. The developed molecular model was employed to design new Cpx poor-clamp and super-clamp mutations and to tested the predictions in silico employing molecular dynamics simulations. Subsequently, we generated Drosophila lines harboring these mutations and confirmed the poor-clamp and super-clamp phenotypes in vivo. Altogether, these results validate the atomic model of the Cpx-mediated fusion clamp, wherein the Cpx AH inserts between the SNARE bundle and the SV lipid bilayer, simultaneously binding the unraveled C terminus of Syb and preventing full SNARE assembly.

Genome-wide identification and characterisation of phenylalanine ammonia-lyase gene family in grapevine

ABSTRACT Phenylalanine ammonia-lyase (PAL) is a rate-limiting enzyme of phenylpropanoid biosynthesis in plants that synthesise many ubiquitous key compounds, including phenolic acids, flavonoids, lignins, and stilbenes. The first draft of the full grape genome assembly was released, but systematic analysis of the PAL family has not been completed in detail. In this study, 15 PALs were identified in the grape genome. Grape PALs are distributed on 6 chromosomes, and 9 PALs are distributed on chromosome 16, in which most of the 2 downstream genes, including chalcone synthase (CHS) and stilbene synthase (STS). Promoter analysis revealed that there were many cis-acting elements in the promoter regions to respond to light, phytohormone, and stress. According to transcriptome-data analysis, VvPALs have extensive expression patterns in various tissues and developmental stages in response to various stresses. Dynamic expression patterns of PAL1/2/3/5 were observed in different developmental stages of 4 grape varieties on the basis of qRT-PCR analysis, and suggested that PALs are involved in a complex mechanism to modulate in anthocyanin biosynthesis. Comprehensive analysis of PALs offers a basis for further understanding the physiological functions of PALs during grape development and their potential role to respond the various stimuli.

A novel canis lupus familiaris reference genome improves variant resolution for use in breed-specific GWAS.

Reference genome fidelity is critically important for genome wide association studies, yet most vary widely from the study population. A typical whole genome sequencing approach implies short-read technologies resulting in fragmented assemblies with regions of ambiguity. Further information is lost by economic necessity when genotyping populations, as lower resolution technologies such as genotyping arrays are commonly used. Here, we present a phased reference genome for Canis lupus familiaris using high molecular weight DNA-sequencing technologies. We tested wet laboratory and bioinformatic approaches to demonstrate a minimum workflow to generate the 2.4 gigabase genome for a Labrador Retriever. The de novo assembly required eight Oxford Nanopore R9.4 flowcells (∼23X depth) and running a 10X Genomics library on the equivalent of one lane of an Illumina NovaSeq S1 flowcell (∼88X depth), bringing the cost of generating a nearly complete reference genome to less than $10K (USD). Mapping of short-read data from 10 Labrador Retrievers against this reference resulted in 1% more aligned reads versus the current reference (CanFam3.1, P < 0.001), and a 15% reduction of variant calls, increasing the chance of identifying true, low-effect size variants in a genome-wide association studies. We believe that by incorporating the cost to produce a full genome assembly into any large-scale genotyping project, an investigator can improve study power, decrease costs, and optimize the overall scientific value of their study.

Evidence for galaxy assembly bias in BOSS CMASS redshift-space galaxy correlation function

ABSTRACT Building accurate and flexible galaxy–halo connection models is crucial in modelling galaxy clustering on non-linear scales. Recent studies have found that halo concentration by itself cannot capture the full galaxy assembly bias effect and that the local environment of the halo can be an excellent indicator of galaxy assembly bias. In this paper, we propose an extended halo occupation distribution (HOD) model that includes both a concentration-based assembly bias term and an environment-based assembly bias term. We use this model to achieve a good fit (χ2/degrees of freedom = 1.35) on the 2D redshift-space two-point correlation function (2PCF) of the Baryon Oscillation Spectroscopic Survey (BOSS) CMASS galaxy sample. We find that the inclusion of both assembly bias terms is strongly favoured by the data and the standard five-parameter HOD model is strongly rejected. More interestingly, the redshift-space 2PCF drives the assembly bias parameters in a way that preferentially assigns galaxies to lower mass haloes. This results in galaxy–galaxy lensing predictions that are within 1σ agreement with the observation, alleviating the perceived tension between galaxy clustering and lensing. We also showcase a consistent 3σ–5σ preference for a positive environment-based assembly bias that persists over variations in the fit. We speculate that the environmental dependence might be driven by underlying processes such as mergers and feedback, but might also be indicative of a larger halo boundaries such as the splashback radius. Regardless, this work highlights the importance of building flexible galaxy–halo connection models and demonstrates the extra constraining power of the redshift-space 2PCF.

Effect of Bird Yaw/Pitch Angles on Soft Impact Damage of a Fan Assembly

This paper presents a numerical investigation of bird attitude angles affecting the soft-impact damage of a full fan assembly. Firstly, considering the geometry of a mallard, a real bird model is established by the Smoothed Particle Hydrodynamics (SPH) method and calibrated with available test data. Then, complying with airworthiness requirements, simulations of a full-bladed fan assembly subjected to a real bird were conducted to determine the critical ingestion parameters (CIP). Furthermore, a real bird with different attitude angles aimed at a full fan assembly was simulated. Results show that attitude angles of the bird produce a significant impact on the effect of the bird strike on rotating blades and would increase the possibility of blade failures, especially for the yaw angle of -45° and the pitch angle of −60°. It is invaluable for commercial airlines and engine manufactures to provide safe flight and landing by adopting the real bird model with critical yaw and pitch angles in the design for resistance to bird ingestion.