关键词： Cottonseed protein concentrate   Largemouth bass   Paraprobiotic   Gut health   Gut microbiome  

摘要： Cottonseed protein concentrate (CPC) is a potential fishmeal alternative but may negatively influence the gut health of fish. A specific paraprobiotic consisting of multi-strain yeast fractions (MsYF) was recently shown to improve gut health via immune modulation. This study aimed to evaluate the effects of MsYF on the growth performance, gut health and microbiome of largemouth bass fed a diet using CPC as 40% fishmeal alternative. Juvenile largemouth bass (body-weight = 31 g) were fed for 65 days with three iso-nitrogen and isoenergetic experimental diets: 1/ one Control diet (50% fishmeal), 2/ one low fishmeal diet (LFM;30% fishmeal, 23.5% CPC) and 3/ one LFM diet + MsYF at 800 mg/kg (LFMY). The highest growth performance, estimated as specific growth rate and protein efficiency ratio, was observed in the LFMY followed by the LFM. Histopathology identified some specimens with distal intestinal lesions in the Control and LFM groups but not of the LFMY group. The gene expression level of il-10 (anti-inflammatory) was up-regulated in the LFM and LFMY groups but only LFMY down-regulated il-1 beta (pro-inflammatory) and hypoxia-inducible factor 1-alpha (hif1 alpha) protein expression level compared to the Control group. Further, LFM and LFMY up-regulated occ-1b (tight-junction) and slc15a2 (peptide transporter) suggesting enhanced intestinal permeability and peptide transport compared to the Control group. Moreover, dietary CPC increased the abundance of genus Clostridium_sensu_stricto_12 and decreased the abundance of genus Escherichia-Shigella compared to the Control group, while the inclusion of MsYF increased the abundance of the potentially beneficial genus Cetobacterium compared to the LFM group. Our results showed 40% substitution of fishmeal protein by CPC could enhance growth performance and regulate the intestinal barrier and peptide transport relevant functions in largemouth bass. Moreover, in a low fishmeal basal diet, MsYF supplementation could increase gr

关键词： calcium   alternative splicing   embryo   protein isoforms   amino acids   exons   skeletal muscles   cell lines   tissue membrane   calcium channels   alanine   sensor   voltage gated calcium channel  

摘要： The skeletal muscle voltage-gated calcium channel (Ca(v)1.1) primarily functions as a voltage sensor for excitation-contraction coupling. Conversely, its ion-conducting function is modulated by multiple mechanisms within the pore-forming a is subunit and the auxiliary alpha(2)delta-1 and gamma(1) subunits. In particular, developmentally regulated alternative splicing of exon 29, which inserts 19 amino acids in the extracellular IVS3-S4 loop of Ca(v)1.1a, greatly reduces the current density and shifts the voltage dependence of activation to positive potentials outside the physiological range. We generated new HEK293 cell lines stably expressing alpha(2)delta-1, beta(3), and STAC3. When the adult (Ca(v)1.1a) and embryonic (Ca(v)1.1e) splice variants were expressed in these cells, the difference in the voltage dependence of activation observed in muscle cells was reproduced, but not the reduced current density of Ca(v)1.1a. Only when we further coexpressed the gamma(1) subunit was the current density of Ca(v)1.1a, but not that of Ca(v)1.1e, reduced by >50%. In addition, gamma(1) caused a shift of the voltage dependence of inactivation to negative voltages in both variants. Thus, the current-reducing effect of gamma(1), unlike its effect on inactivation, is specifically dependent on the inclusion of exon 29 in Ca(v)1.1a. Molecular structure modeling revealed several direct ionic interactions between residues in the IVS3-S4 loop and the y i subunit. However, substitution of these residues by alanine, individually or in combination, did not abolish the gamma(1)-dependent reduction of current density, suggesting that structural rearrangements in Ca(v)1.1a induced by inclusion of exon 29 may allosterically empower the gamma(1) subunit to exert its inhibitory action on Ca-v1.1 calcium currents.

摘要： Alternative polyadenylation (APA) plays an important role in gene regulation. With the recent application of novel sequencing technology in APA profiling, an ever-increasing number of APA genes/sites have been identified. However, the phenotypic relevance of most of these APA isoforms remains elusive, which is largely due to the lack of a convenient genetics tool for APA interference. To address this issue, herein, an efficient method is developed based on the CRISPR-dCas13 system, termed as CRISPR-iPAS. Out of eight different dCas13 proteins, Porphyromonas gulae (Pgu) dCas13b, is identified as the most effective one in blocking the usage of the polyadenylation site (PAS). With guide RNAs targeting at core regulatory elements, dPguCas13b enabled APA regulation of endogenous genes with different APA types, including tandem 3 ' UTR, alternative terminal exon, as well as intronic PAS. Finally, we demonstrated that the proposed APA perturbation tool could be used to investigate the functional relevance of APA isoforms.

摘要： Most human protein-coding genes produce alternative polyadenylation (APA) isoforms that differ in 3 ' UTR size or, when coupled with splicing, have variable coding sequences. APA is an important layer of gene expression program critical for defining cell identity. Here, by using a catalytically dead Cas9 and coupling its target site with polyadenylation site (PAS), we develop a method, named CRISPRpas, to alter APA isoform abundance. CRISPRpas functions by enhancing proximal PAS usage, whose efficiency is influenced by several factors, including targeting strand of DNA, distance between PAS and target sequence and strength of the PAS. For intronic polyadenylation (IPA), splicing features, such as strengths of 5 ' splice site and 3 ' splice site, also affect CRISPRpas efficiency. We show modulation of APA of multiple endogenous genes, including IPA of PCF11, a master regulator of APA and gene expression. In sum, CRISPRpas offers a programmable tool for APA regulation that impacts gene expression.

摘要： Ssd1, a conserved fungal RNA-binding protein, is important in stress responses, cell division and virulence. Ssd1 is closely related to Dis3L2 of the RNase II family of nucleases, but lacks catalytic activity and likely suppresses translation of bound mRNAs. Previous studies identified RNA motifs enriched in Ssd1-associated transcripts, yet the sequence requirements for Ssd1 binding are not defined. Here, we identify precise binding sites of Ssd1 on RNA using in vivo cross-linking and cDNA analysis. These sites are enriched in 5 ' untranslated regions of a subset of mRNAs encoding cell wall proteins. We identified a conserved bipartite motif that binds Ssd1 with high affinity in vitro. Active RNase II enzymes have a characteristic, internal RNA binding path;the Ssd1 crystal structure at 1.9 angstrom resolution shows that remnants of regulatory sequences block this path. Instead, RNA binding activity has relocated to a conserved patch on the surface of the protein. Structure-guided mutations of this surface prevent Ssd1 from binding RNA in vitro and phenocopy Ssd1 deletion in vivo. These studies provide a new framework for understanding the function of a pleiotropic post-transcriptional regulator of gene expression and give insights into the evolution of regulatory and binding elements in the RNase II family.

关键词： Spray penetration   spray cone angle   injection pressure   alternative fuels   evaporation model  

摘要： Spray characteristics and transient behaviour of alternative fuel droplets at different spray cone angles, injection pressures and droplet diameters are presented, using a simple spray evaporation model. The model only considers drag force acting on the droplet. Radiation heat transfer during the evaporation process is neglected. The model incorporates temperature-dependent fuel properties, drag coefficient and Reynolds number for robustness. It is tested for alternative fuels, namely Jatropha Bio-synthetic Paraffinic Kerosen (JSPK), Camelina Bio-synthetic Paraffinic Kerosene (CSPK) and biodiesel, while Jet-A is used for comparison. The transient behaviour of fuel droplets is visualised through a time-history of the normalised squared diameter. Changes in the particles' temperature, mass, density, velocity and heat transfer coefficient over time are observed. Moreover, penetration of the fuels at different spray cone angles, injection pressures and droplet diameters is compared. Among the tested fuels, biodiesel is found to have the farthest penetration due to its high vapour pressure. Notably, increasing spray cone angle reduces penetration length. A similar trend is observed when injection pressure is increased. However, at constant injection pressure, increasing droplet diameter increases spray cone angle and spray penetration. To obtain optimum penetration, therefore, a larger cone angle should be used for fuel with high density and vapour pressure, and vice versa.

摘要： 莫言有一种能力,就是非常有效地将现实生活转化为非现实生活,没有比他小说里的现实生活更不现实了.仿佛他看世界的眼睛有一种屈光功能,景物一旦进入视野,顿时就改了面目.并不是说与原来完全不一样,甚至很一样,可就是成了另一个世界.这世界里的一切还是依原来的样式链接镶嵌,色彩却全变了,很容易将其视为一种风格,但风格其实是装饰的意味,这里的色彩则影响到事情的性质.这"色彩"更接近"质地"的意思,事情的质地不同了,于是,就变得不那么真实.在那个不真实的世界里,比真实的更为沉重,但确有着一重意境.小说实在是一种过于结实的东西,现实既是它的内核,又是外相,要从中抽离出一个独立的世界,是需要更有力量的占位.

关键词： Ozone   Decontamination   SARS-CoV-2   Protective equipment   COVID-19  

摘要： COVID-19 pandemic created a global shortage of medical protective equipment. Here, we considered ozone (O3) a disinfectant alternative due to its potent oxidative activity against biological macromolecules. The O3 decontamination assays were done using SARS-CoV-2 obtained from patients to produce artificial contamination of N95 masks and biosecurity gowns. The quantification of SARS-CoV-2 was performed before and after exposing the samples to different ozone gas concentrations for times between 5 and 30 min. Viral loads as a function of the O3 exposure time were estimated from the data obtained by the RT-PCR technique. The genetic material of the virus was no longer detected for any tested concentrations after 15 min of O3 exposure, which means a disinfection Concentration-Time above 144 ppm min. Vibrational spectroscopies were used to follow the modifications of the polymeric fibers after the O3 treatment. The results indicate that the N95 masks could be safely reused after decontamination with treatments of 15 min at the established O3 doses for a maximum of 6 cycles.

关键词： Land-use conflict   Energy system   Modeling to generate alternatives   Uncertainty   Energy transition  

摘要： Long-term power market outlooks suggest a rapid increase in renewable energy deployment as a main solution to greenhouse gas mitigation in the Northern European energy system. However, the consequential area requirement is a non-techno-economic aspect that currently is omitted by many energy system optimization models. This study applies modeling to generate alternatives (MGA) technique to the Balmorel energy system model to address spatial conflicts related to increased renewable energy deployment. The approach searches for alternative solutions that minimize land-use conflicts while meeting the low-carbon target, by allowing a 1% to 15% increase in system costs compared to the least-cost solution. Two alternative objectives are defined to reflect various aspects of spatial impact. The results show that the least-cost solution requires 1.2%-3.6% of the land in the modeled countries in 2040 for onshore wind and solar PV installations. A 10% increase in costs can reduce the required land area by 58% by relying more on offshore wind. Nuclear energy may also be an option if both onshore and offshore areas are to be reduced, or in a less flexible system. Both offshore wind and nuclear energy technologies are associated with higher risks and pose uncertainties in terms of reaching the climate targets in time. The changes in costs and required land areas imply significantly higher annual costs ranging from 200 to 750 kEUR/km2 to avoid land use for energy infrastructure. Overall, this study confirms that the energy transition strategies prioritizing land savings from energy infrastructure are feasible, but high risks and costs of averted land are involved.