The tetraplex (CGG)n destabilizing proteins hnRNP A2 and CBF-A enhance the in vivo translation of fragile X premutation mRNA

Expansion of a (CGG)n sequence in the 5′-UTR of the FMR1 gene to >200–2000 repeats abolishes its transcription and initiates fragile X syndrome (FXS). By contrast, levels of FMR1 mRNA are 5–10-fold higher in FXS premutation carriers of >55–200 repeats than in normal subjects. Lack of a corresponding increase in the amount of the product FMRP protein in carrier cells suggest that (CGG)>55–200 tracts thwart translation. Here we report that a (CGG)99 sequence positioned upstream to reporter firefly (FL) gene selectively diminished mRNA translation in coupled and separate T7 promoter-driven in vitro transcription and translation systems. The (CGG)99 tract similarly depressed mRNA utilization in HEK293 human cells transfected with plasmids bearing FMR1 promoter-driven FL gene. A (CGG)33 RNA tract formed a largely RNase T1-resistant intramolecular secondary structure in the presence of K+ ions. Expression of the quadruplex (CGG)n disrupting proteins hnRNP A2 or CBF-A in HEK293 cells significantly elevated the efficacy of (CGG)99 FL mRNA translation whereas hnRNP A2 or CBF-A mutants lacking quadruplex (CGG)n disrupting activity did not. Taken together, our results suggest that secondary structures of (CGG)n in mRNA obstruct its translation and that quadruplex-disrupting proteins alleviate the translational block

Managing Polyploidy in Ex Situ Conservation Genetics: The Case of the Critically Endangered Adriatic Sturgeon (Acipenser naccarii)

While the current expansion of conservation genetics enables to address more efficiently the management of threatened species, alternative methods for genetic relatedness data analysis in polyploid species are necessary. Within this framework, we present a standardized and simple protocol specifically designed for polyploid species that can facilitate management of genetic diversity, as exemplified by the ex situ conservation program for the tetraploid Adriatic sturgeon Acipenser naccarii. A critically endangered endemic species of the Adriatic Sea tributaries, its persistence is strictly linked to the ex situ conservation of a single captive broodstock currently decimated to about 25 individuals, which represents the last remaining population of Adriatic sturgeon of certain wild origin. The genetic variability of three F1 broodstocks available as future breeders was estimated based on mitochondrial and microsatellite information and compared with the variability of the parental generation. Genetic data showed that the F1 stocks have only retained part of the genetic variation present in the original stock due to the few parent pairs used as founders. This prompts for the urgent improvement of the current F1 stocks by incorporating new founders that better represent the genetic diversity available. Following parental allocation based on band sharing values, we set up a user-friendly tool for selection of candidate breeders according to relatedness between all possible parent-pairs that secures the use of non-related individuals. The approach developed here could also be applied to other endangered tetraploid sturgeon species overexploited for caviar production, particularly in regions lacking proper infrastructure and/or expertise

Cool, Luminous, and Highly Variable Stars in the Magellanic Clouds. II: Spectroscopic and Environmental Analysis of Thorne-\.Zytkow Object and Super-AGB Star Candidates

In previous work we identified a population of 38 cool and luminous variable stars in the Magellanic Clouds and examined 11 in detail in order to classify them as either Thorne-\.Zytkow Objects (T\.ZOs, red supergiants with a neutron star cores) or super-AGB stars (the most massive stars that will not undergo core collapse). This population includes HV\,2112, a peculiar star previously considered in other works to be either a T\.ZO or high-mass AGB star. Here we continue this investigation, using the kinematic and radio environments and local star formation history of these stars to place constraints on the age of the progenitor systems and the presence of past supernovae. These stars are not associated with regions of recent star formation, and we find no evidence of past supernovae at their locations. Finally, we also assess the presence of heavy elements and lithium in their spectra compared to red supergiants. We find strong absorption in Li and s-process elements compared to RSGs in most of the sample, consistent with super-AGB nucleosynthesis, while HV\,2112 shows additional strong lines associated with T\.ZO nucleosynthesis. Coupled with our previous mass estimates, the results are consistent with the stars being massive (~4-6.5M$_{\odot}$) or super-AGB (~6.5-12M$_{\odot}$) stars in the thermally pulsing phase, providing crucial observations of the transition between low- and high-mass stellar populations. HV\,2112 is more ambiguous; it could either be a maximally massive sAGB star, or a T\.ZO if the minimum mass for stability extends down to <13 M$_\odot$.Comment: 32 pages, 10 figures, 7 tables, spectroscopic data available at https://zenodo.org/record/7058608, accepted to The Astrophysical Journa

Exploring functional pairing between surface glycoconjugates and human galectins using programmable glycodendrimersomes

Precise translation of glycan-encoded information into cellular activity depends critically on highly specific functional pairing between glycans and their human lectin counter receptors. Sulfoglycolipids, such as sulfatides, are important glycolipid components of the biological membranes found in the nervous and immune systems. The optimal molecular and spatial design aspects of sulfated and nonsulfated glycans with high specificity for lectin-mediated bridging are unknown. To elucidate how different molecular and spatial aspects combine to ensure the high specificity of lectin-mediated bridging, a bottom-up toolbox is devised. To this end, negatively surface-charged glycodendrimersomes (GDSs), of different nanoscale dimensions, containing sulfo-lactose groups are self-assembled in buffer from a synthetic sulfatide mimic: Janus glycodendrimer (JGD) containing a 3′-O-sulfo-lactose headgroup. Also prepared for comparative analysis are GDSs with nonsulfated lactose, a common epitope of human membranes. These self-assembled GDSs are employed in aggregation assays with 15 galectins, comprising disease-related human galectins, and other natural and engineered variants from four families, having homodimeric, heterodimeric, and chimera architectures. There are pronounced differences in aggregation capacity between human homodimeric and heterodimeric galectins, and also with respect to their responsiveness to the charge of carbohydrate-derived ligand. Assays reveal strong differential impact of ligand surface charge and density, as well as lectin concentration and structure, on the extent of surface cross-linking. These findings demonstrate how synthetic JGD-headgroup tailoring teamed with protein engineering and network assays can help explain how molecular matchmaking operates in the cellular context of glycan and lectin complexity

Development of a Fully Human Anti-PDGFRβ Antibody That Suppresses Growth of Human Tumor Xenografts and Enhances Antitumor Activity of an Anti-VEGFR2 Antibody

Platelet-derived growth factor receptor β (PDGFRβ) is upregulated in most of solid tumors. It is expressed by pericytes/smooth muscle cells, fibroblast, macrophage, and certain tumor cells. Several PDGF receptor-related antagonists are being developed as potential antitumor agents and have demonstrated promising antitumor activity in both preclinical and clinical settings. Here, we produced a fully human neutralizing antibody, IMC-2C5, directed against PDGFRβ from an antibody phage display library. IMC-2C5 binds to both human and mouse PDGFRβ and blocks PDGF-B from binding to the receptor. IMC-2C5 also blocks ligand-stimulated activation of PDGFRβ and downstream signaling molecules in tumor cells. In animal studies, IMC-2C5 significantly delayed the growth of OVCAR-8 and NCI-H460 human tumor xenografts in nude mice but failed to show antitumor activities in OVCAR-5 and Caki-1 xenografts. Our results indicate that the antitumor efficacy of IMC-2C5 is primarily due to its effects on tumor stroma, rather than on tumor cells directly. Combination of IMC-2C5 and DC101, an anti-mouse vascular endothelial growth factor receptor 2 antibody, resulted in significantly enhanced antitumor activity in BxPC-3, NCI-H460, and HCT-116 xenografts, compared with DC101 alone, and the trend of additive effects to DC101 treatment in several other tumor models. ELISA analysis of NCI-H460 tumor homogenates showed that IMC-2C5 attenuated protein level of vascular endothelial growth factor and basic fibroblast growth factor elevated by DC101 treatment. Finally, IMC-2C5 showed a trend of additive effects when combined with DC101/chemotherapy in MIA-PaCa-2 and NCI-H460 models. Taken together, these results lend great support to the use of PDGFRβ antagonists in combination with other antiangiogenic agents in the treatment of a broad range of human cancers

Sexual Dimorphic Regulation of Body Weight Dynamics and Adipose Tissue Lipolysis

BACKGROUND: Successful reduction of body weight (BW) is often followed by recidivism to obesity. BW-changes including BW-loss and -regain is associated with marked alterations in energy expenditure (EE) and adipose tissue (AT) metabolism. Since these processes are sex-specifically controlled, we investigated sexual dimorphisms in metabolic processes during BW-dynamics (gain-loss-regain). RESEARCH DESIGN: Obesity was induced in C57BL/6J male (m) and female (f) mice by 15 weeks high-fat diet (HFD) feeding. Subsequently BW was reduced (-20%) by caloric restriction (CR) followed by adaptive feeding, and a regain-phase. Measurement of EE, body composition, blood/organ sampling were performed after each feeding period. Lipolysis was analyzed ex-vivo in gonadal AT. RESULTS: Male mice exhibited accelerated BW-gain compared to females (relative BW-gain m:140.5±3.2%; f:103.7±6.5%; p<0.001). In consonance, lean mass-specific EE was significantly higher in females compared to males during BW-gain. Under CR female mice reached their target-BW significantly faster than male mice (m:12.2 days; f:7.6 days; p<0.001) accompanied by a sustained sex-difference in EE. In addition, female mice predominantly downsized gonadal AT whereas the relation between gonadal and total body fat was not altered in males. Accordingly, only females exhibited an increased rate of forskolin-stimulated lipolysis in AT associated with significantly higher glycerol concentrations, lower RER-values, and increased AT expression of adipose triglyceride lipase (ATGL) and hormone sensitive lipase (HSL). Analysis of AT lipolysis in estrogen receptor alpha (ERα)-deficient mice revealed a reduced lipolytic rate in the absence of ERα exclusively in females. Finally, re-feeding caused BW-regain faster in males than in females. CONCLUSION: The present study shows sex-specific dynamics during BW-gain-loss-regain. Female mice responded to CR with an increase in lipolytic activity, and augmented lipid-oxidation leading to more efficient weight loss. These processes likely involve ERα-dependent signaling in AT and sexual dimorphic regulation of genes involved in lipid metabolism

Survey of Period Variations of Superhumps in SU UMa-Type Dwarf Novae. VIII: The Eighth Year (2015-2016)

Continuing the project described by Kato et al. (2009, arXiv:0905.1757), we collected times of superhump maxima for 128 SU UMa-type dwarf novae observed mainly during the 2015-2016 season and characterized these objects. The data have improved the distribution of orbital periods, the relation between the orbital period and the variation of superhumps, the relation between period variations and the rebrightening type in WZ Sge-type objects. Coupled with new measurements of mass ratios using growing stages of superhumps, we now have a clearer and statistically greatly improved evolutionary path near the terminal stage of evolution of cataclysmic variables. Three objects (V452 Cas, KK Tel, ASASSN-15cl) appear to have slowly growing superhumps, which is proposed to reflect the slow growth of the 3:1 resonance near the stability border. ASASSN-15sl, ASASSN-15ux, SDSS J074859.55+312512.6 and CRTS J200331.3-284941 are newly identified eclipsing SU UMa-type (or WZ Sge-type) dwarf novae. ASASSN-15cy has a short (~0.050 d) superhump period and appears to belong to EI Psc-type objects with compact secondaries having an evolved core. ASASSN-15gn, ASASSN-15hn, ASASSN-15kh and ASASSN-16bu are candidate period bouncers with superhump periods longer than 0.06 d. We have newly obtained superhump periods for 79 objects and 13 orbital periods, including periods from early superhumps. In order that the future observations will be more astrophysically beneficial and rewarding to observers, we propose guidelines how to organize observations of various superoutbursts.Comment: 123 pages, 162 figures, 119 tables, accepted for publication in PASJ (including supplementary information

Accommodation and vergence response gains to different near cues characterize specific esotropias

Aim. To describe preliminary findings of how the profile of the use of blur, disparity and proximal cues varies between non-strabismic groups and those with different types of esotropia. Design. Case control study Methodology. A remote haploscopic photorefractor measured simultaneous convergence and accommodation to a range of targets containing all combinations of binocular disparity, blur and proximal (looming) cues. 13 constant esotropes, 16 fully accommodative esotropes, and 8 convergence excess esotropes were compared with age and refractive error matched controls, and 27 young adult emmetropic controls. All wore full refractive correction if not emmetropic. Response AC/A and CA/C ratios were also assessed. Results. Cue use differed between the groups. Even esotropes with constant suppression and no binocular vision (BV) responded to disparity in cues. The constant esotropes with weak BV showed trends for more stable responses and better vergence and accommodation than those without any BV. The accommodative esotropes made less use of disparity cues to drive accommodation (p=0.04) and more use of blur to drive vergence (p=0.008) than controls. All esotropic groups failed to show the strong bias for better responses to disparity cues found in the controls, with convergence excess esotropes favoring blur cues. AC/A and CA/C ratios existed in an inverse relationship in the different groups. Accommodative lag of >1.0D at 33cm was common (46%) in the pooled esotropia groups compared with 11% in typical children (p=0.05). Conclusion. Esotropic children use near cues differently from matched non-esotropic children in ways characteristic to their deviations. Relatively higher weighting for blur cues was found in accommodative esotropia compared to matched controls

SPR imaging biosensor for the 20S proteasome: sensor development and application to measurement of proteasomes in human blood plasma

The 20S proteasome is a multicatalytic enzyme complex responsible for intracellular protein degradation in mammalian cells. Its antigen level or enzymatic activity in blood plasma are potentially useful markers for various malignant and nonmalignant diseases. We have developed a method for highly selective determination of the 20S proteasome using a Surface Plasmon Resonance Imaging (SPRI) technique. It is based on the highly selective interaction between the proteasome’s catalytic β5 subunit and immobilized inhibitors (the synthetic peptide PSI and epoxomicin). Inhibitor concentration and pH were optimized. Analytical responses, linear ranges, accuracy, precision and interferences were investigated. Biosensors based on either PSI and epoxomicin were found to be suitable for quantitative determination of the proteasome, with a precision of ±10% for each, and recoveries of 102% and 113%, respectively, and with little interference by albumin, trypsin, chymotrypsin, cathepsin B and papain. The proteasome also was determined in plasma of healthy subjects and of patients suffering from acute leukemia. Both biosensors gave comparable results (2860 ng·mL-1 on average for control, and 42300 ng·mL-1 on average for leukemia patients)

Postprandial differences in the plasma metabolome of healthy Finnish subjects after intake of a sourdough fermented endosperm rye bread versus white wheat bread

<p>Abstract</p> <p>Background</p> <p>The mechanism behind the lowered postprandial insulin demand observed after rye bread intake compared to wheat bread is unknown. The aim of this study was to use the metabolomics approach to identify potential metabolites related to amino acid metabolism involved in this mechanism.</p> <p>Methods</p> <p>A sourdough fermented endosperm rye bread (RB) and a standard white wheat bread (WB) as a reference were served in random order to 16 healthy subjects. Test bread portions contained 50 g available carbohydrate. <it>In vitro </it>hydrolysis of starch and protein were performed for both test breads. Blood samples for measuring glucose and insulin concentrations were drawn over 4 h and gastric emptying rate (GER) was measured. Changes in the plasma metabolome were investigated by applying a comprehensive two-dimensional gas chromatography coupled to time-of-flight mass spectrometry metabolomics platform (GC×GC-TOF-MS).</p> <p>Results</p> <p>Plasma insulin response to RB was lower than to WB at 30 min (P = 0.004), 45 min (P = 0.002) and 60 min (P < 0.001) after bread intake, and plasma glucose response was significantly higher at time point 90 min after RB than WB intake (P = 0.045). The starch hydrolysis rate was higher for RB than WB, contrary to the <it>in vitro </it>protein digestibility. There were no differences in GER between breads. From 255 metabolites identified by the metabolomics platform, 26 showed significant postprandial relative changes after 30 minutes of bread intake (p and q values < 0.05). Among them, there were changes in essential amino acids (phenylalanine, methionine, tyrosine and glutamic acid), metabolites involved in the tricarboxylic acid cycle (alpha-ketoglutaric, pyruvic acid and citric acid) and several organic acids. Interestingly, the levels of two compounds involved in the tryptophan metabolism (picolinic acid, ribitol) significantly changed depending on the different bread intake.</p> <p>Conclusions</p> <p>A single meal of a low fibre sourdough rye bread producing low postprandial insulin response brings in several changes in plasma amino acids and their metabolites and some of these might have properties beneficial for health.</p