As an ATP-dependent DNA helicase, human ChlR1/DDX11 (Chl1 in yeast) can unwind both DNA:RNA and DNA:DNA substrates in vitro. Studies have demonstrated that ChlR1 plays a vital role in preserving ...genome stability by participating in DNA repair and sister chromatid cohesion, whereas the ways in which the biochemical features of ChlR1 function in DNA metabolism are not well understood. Here, we illustrate that Chl1 localizes to double-strand DNA break (DSB) sites and restrains DNA:RNA hybrid accumulation at these loci. Mutation of Chl1 strongly impairs DSB repair capacity by homologous recombination (HR) and nonhomologous end-joining (NHEJ) pathways, and deleting RNase H further reduces DNA repair efficiency, which indicates that the enzymatic activities of Chl1 are needed in Schizosaccharomyces pombe. In addition, we found that the Rpc37 subunit of RNA polymerase III (RNA Pol III) interacts directly with Chl1 and that deletion of Chl1 has no influence on the localization of Rpc37 at DSB site, implying the role of Rpc37 in the recruitment of Chl1 to this site.
•Resin methods were employed to remove the proteins and pigments from polysaccharides of Gentiana scabra Bunge roots.•The weight-average molecular weights of purified polysaccharides (named GSP-2 and ...GSP-3) were analyzed by HPGPC.•Monosaccharide compositions of polysaccharide fractions were measured by HPAEC-PAD.•GSP-3 showed significant anticoagulant activity in APTT and TT assays.
Three water-soluble polysaccharide fractions (GSP-1, GSP-2 and GSP-3) were obtained from Gentiana scabra Bunge roots by DEAE-Sepharose CL-6B and Sepharose CL-6B column chromatography. Their chemical characterizations were determined by high performance gel permeation chromatography (HPGPC), high performance anion exchange chromatography coupled with pulsed amperometric detection (HPAEC-PAD) and Fourier transform infrared (FT-IR) spectrometer. Moreover, their in vitro anticoagulant activities were evaluated by activated partial thromboplastin time (APTT), thrombin time (TT) and prothrombin time (PT) assays. GSP-1 and GSP-2 were composed of rhamnose, arabinose, galactose, glucose and galacturonic acid, while GSP-3 consisted of rhamnose, arabinose, galactose and galacturonic acid with a weight-average molecular weight of 5.8×104Da. In comparison with the control group (saline), GSP, GSP-1, GSP-2 and GSP-3 could prolong APTT and TT, but not PT. Overall, GSP-3 exhibited potent anticoagulant activity and would be expected to be a potential source of anticoagulant.
This paper aims at designing a distributed static output feedback (DSOF) controller for the interconnected system in finite‐frequency domain. Motivated by the fact that the signal transmitting ...process is usually affected by surrounding environments, the interconnection communication between subsystems are assumed to be non‐ideal. By introducing some dilated multipliers into the extended condition that guarantees the well‐posedness, stability and finite‐frequency H∞ performance of non‐ideally interconnected systems, a new equivalent condition is derived, which is more effective for parameterizing the DSOF controller. A two‐stage approach is then developed for constructing a DSOF controller: design an initial distributed full information (DFI) controller first and then derive a desired DSOF from the DFI controller. Moreover, two iterative linear matrix inequality based algorithms are proposed to improve the solvability of the DFI controller and DSOF controller design problems. Finally, an example is given to show the effectiveness of the proposed two‐stage DSOF controller design method.
Outbreaks of Classical swine fever virus (CSFV) cause significant economic losses in the swine industry. Vaccination is the major method to prevent and control the disease. As live attenuated ...vaccines fail to elicit differentiable immunity between infected and vaccinated animals, subunit vaccine was considered as an alternative candidate to prevent and eradicate CSFV. Subunit vaccines present advantages in DIVA immunogenicity and safety. The technology was limited due to the low yield and the high cost with multiple and large doses. The native E2 signal peptide has not been well defined before. Here, the aim of this study is to develop a cost-effective and efficacious E2 vaccine candidate against CSFV with signal peptide and E2 sequence selection.
A novel CSFV E2 sequence (E2ZJ) was identified from an epidemic strain of Zhejiang for outstanding secretion in baculovirus and enhanced immunogenicity. E2 secretion induced with the selected signal peptide, SPZJ (SP23), increase at least 50% as compared to any other signal peptides tested. Besides, unique antigenic features were identified in E2ZJ. As indicated with immunized sera in IFA against CSFV infection, E2ZJ elicited CSFV antibodies at the earlier stage than other E2 types tested in mice. Moreover, higher level of neutralizing and CSFV antibodies against CSFV with E2ZJ was detected than other E2s with the same dosage at 28 dpi. Further, E2ZJ successfully elicited neutralizing immunity in piglets. A single dose of 5 μg of E2ZJ was sufficient to induce protective antibodies against CSFV in piglets and provided 100% protection against lethal virus challenge.
Our studies provide evidence that E2ZJ guided by a novel E2 signal peptide (SPZJ) was efficiently secreted and presented significantly improved immunogenicity than conventional E2 vaccines. Moreover, a single dose of 5 μg E2ZJ is efficacious against CSFV in piglets.
With the economic rise of China, there is global demand for effective teaching and learning of Chinese as a foreign language (CFL). There has been limited sustained success in Chinese language ...learning in Australian schools, however, and this has been attributed, amongst other factors, to pedagogy employed by teachers. Today, it is commonplace to understand that educational background influences teacher beliefs which in turn impact pedagogical transition in overseas teaching environments. This exploratory case study reports qualitative analysis of interviews with nine school teachers of CFL in New South Wales, Australia. The study mapped three groups of beliefs within the Chinese teacher group, namely, beliefs which align with principles of traditional Chinese education, beliefs which align with constructivist learning principles and beliefs in transition between. All teachers in the CFL teacher community of practice could be said to be in a fluid process of transition, to find new pedagogical identities and best practice. This study's findings as to transitioning beliefs, and the study's recommendations, are of significance in the design of more effective teacher training suitable to achieve successful learning outcomes in Chinese foreign language classrooms.
GATA1-related cytopenia (GRC) is characterized by thrombocytopaenia and/or anaemia ranging from mild to severe. Haematopoietic stem cell transplantation (HSCT) is a healing therapeutic choice for GRC ...patients. We identified a novel pathogenic variant (GATA1: c.1019delG) in a boy with GATA1-related cytopenia. Then we performed preimplantation genetic testing (PGT) in this GRC family. After a mosaic embryo transfered, a healthy and HLA-compatible with the proband baby was delivered. NGS-based PGT-HLA is a valuable procedure for the treatment of GATA1-related cytopenia caused by GATA1 variants, or other haematological disorders, oncological and immunological diseases. Furthermore, our study reconfirms that mosaic embryos transfer would bring healthy offspring.
Meiotic crossovers are produced when programmed double-strand breaks (DSBs) are repaired by recombination from homologous chromosomes (homologues). In a wide variety of organisms, meiotic ...HORMA-domain proteins are required to direct DSB repair towards homologues. This inter-homologue bias is required for efficient homology search, homologue alignment, and crossover formation. HORMA-domain proteins are also implicated in other processes related to crossover formation, including DSB formation, inhibition of promiscuous formation of the synaptonemal complex (SC), and the meiotic prophase checkpoint that monitors both DSB processing and SCs. We examined the behavior of two previously uncharacterized meiosis-specific mouse HORMA-domain proteins--HORMAD1 and HORMAD2--in wild-type mice and in mutants defective in DSB processing or SC formation. HORMADs are preferentially associated with unsynapsed chromosome axes throughout meiotic prophase. We observe a strong negative correlation between SC formation and presence of HORMADs on axes, and a positive correlation between the presumptive sites of high checkpoint-kinase ATR activity and hyper-accumulation of HORMADs on axes. HORMADs are not depleted from chromosomes in mutants that lack SCs. In contrast, DSB formation and DSB repair are not absolutely required for depletion of HORMADs from synapsed axes. A simple interpretation of these findings is that SC formation directly or indirectly promotes depletion of HORMADs from chromosome axes. We also find that TRIP13 protein is required for reciprocal distribution of HORMADs and the SYCP1/SC-component along chromosome axes. Similarities in mouse and budding yeast meiosis suggest that TRIP13/Pch2 proteins have a conserved role in establishing mutually exclusive HORMAD-rich and synapsed chromatin domains in both mouse and yeast. Taken together, our observations raise the possibility that involvement of meiotic HORMA-domain proteins in the regulation of homologue interactions is conserved in mammals.
Ribosomally synthesized and post-translationally modified peptides (RiPPs) are a major class of natural products with diverse chemical structures and potent biological activities. A vast majority of ...RiPP gene clusters remain unexplored in microbial genomes, which is partially due to the lack of rapid and efficient heterologous expression systems for RiPP characterization and biosynthesis. Here, we report a unified biocatalysis (UniBioCat) system based on cell-free gene expression for rapid biosynthesis and engineering of RiPPs. We demonstrate UniBioCat by reconstituting a full biosynthetic pathway for de novo biosynthesis of salivaricin B, a lanthipeptide RiPP. Next, we delete several protease/peptidase genes from the source strain to enhance the performance of UniBioCat, which then can synthesize and screen salivaricin B variants with enhanced antimicrobial activity. Finally, we show that UniBioCat is generalizable by synthesizing and evaluating the bioactivity of ten uncharacterized lanthipeptides. We expect UniBioCat to accelerate the discovery, characterization, and synthesis of RiPPs.
The cohesin protein complex contributes to transcriptional regulation in a CTCF-independent manner by colocalizing with master regulators at tissue-specific loci. The regulation of transcription ...involves the concerted action of multiple transcription factors (TFs) and cohesin's role in this context of combinatorial TF binding remains unexplored. To investigate cohesin-non-CTCF (CNC) binding events in vivo we mapped cohesin and CTCF, as well as a collection of tissue-specific and ubiquitous transcriptional regulators using ChIP-seq in primary mouse liver. We observe a positive correlation between the number of distinct TFs bound and the presence of CNC sites. In contrast to regions of the genome where cohesin and CTCF colocalize, CNC sites coincide with the binding of master regulators and enhancer-markers and are significantly associated with liver-specific expressed genes. We also show that cohesin presence partially explains the commonly observed discrepancy between TF motif score and ChIP signal. Evidence from these statistical analyses in wild-type cells, and comparisons to maps of TF binding in Rad21-cohesin haploinsufficient mouse liver, suggests that cohesin helps to stabilize large protein-DNA complexes. Finally, we observe that the presence of mirrored CTCF binding events at promoters and their nearby cohesin-bound enhancers is associated with elevated expression levels.
This study develops a mixed behavioural equilibrium model with explicit consideration of mode choice (MBE-MC) in a transportation system where fully automated vehicles (AV) coexist with conventional ...human-driven vehicles (HV). For the mode choice, travellers select among three options, following a logit modal split: driving their private HV, or taking an AV mobility service provided by either a firm or the government. For the route choice, the HV drivers follow the random utility maximisation principle while central agents route the AV passengers following the Cournot Nash (firm agent) or Social Optimal (government agent) principles. We consider two types of travel costs (i.e. travel time and monetary travel cost) to characterise the new features (e.g. expanded link capacity and reduced value of time) of the mixed AV–HV transportation system. We model the MBE-MC problem in a combined mode–route choice framework and formulate it as a route-based variational inequality (VI) problem. We show the equivalence between the VI formulation and the MBE-MC problem, and the existence of a solution to the MBE-MC problem. Then, we modify a partial linearisation algorithm for solving the proposed model. Numerical results validate the equilibrium conditions and show the efficacy of the new model in capturing the features of the mixed AV–HV transportation system. The impact patterns of different parameters on (1) the network performance in terms of AV share and system cost and (2) on the solution efficiency are analysed.
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