Molecular basis of the dominant negative effect of a glycine transporter 2 mutation associated with hyperekplexia

"This research was originally published in Journal of Biological Chemistry. Esther Arribas-González, Jaime de Juan-Sanz, Carmen Aragón, and Beatriz López-Corcuera. Molecular Basis of the Dominant Negative Effect of a Glycine Transporter 2 Mutation Associated with Hyperekplexia. Journal of Biological Chemistry. 2015; 290:2150-2165. © the American Society for Biochemistry and Molecular Biology."Hyperekplexia or startle disease is a rare clinical syndrome characterized by an exaggerated startle in response to trivial tactile or acoustic stimuli. This neurological disorder can have serious consequences in neonates, provoking brain damage and/or sudden death due to apnea episodes and cardiorespiratory failure. Hyperekplexia is caused by defective inhibitory glycinergic neurotransmission. Mutations in the human SLC6A5 gene encoding the neuronal GlyT2 glycine transporter are responsible for the presynaptic form of the disease. GlyT2 mediates synaptic glycine recycling, which constitutes the main source of releasable transmitter at glycinergic synapses. Although the majority of GlyT2 mutations detected so far are recessive, a dominant negative mutant that affects GlyT2 trafficking does exist. In this study, we explore the properties and structural alterations of the S512R mutation in GlyT2. We analyze its dominant negative effect that retains wild-type GlyT2 in the endoplasmic reticulum (ER), preventing surface expression. Weshow that the presence of an arginine rather than serine 512 provoked transporter misfolding, enhanced association to the ER-chaperone calnexin, altered association with the coat-protein complex II component Sec24D, and thereby impeded ER exit. The S512R mutant formed oligomers with wild-type GlyT2 causing its retention in the ER. Overexpression of calnexin rescued wild-type GlyT2 from the dominant negative effect of the mutant, increasing the amount of transporter that reached the plasma membrane and dampening the interaction between the wild-type and mutant GlyT2. The ability of chemical chaperones to overcome the dominant negative effect of the disease mutation on the wild-type transporter was demonstrated in heterologous cells and primary neuronsThis work was supported by Spanish “Ministerio de Economía y Competitividad” Grant SAF2011-28674, by the Centro de Investigación Biomédica en Red de Enfermedades Raras (CIBERER), and by an institutional grant from the “Fundación Ramón Areces.

LENGTH-WEIGHT RELATIONSHIPS FOR SEVERAL LARGE PELAGIC SHARKS FROM THE INDIAN OCEAN

Fork length-dressed weight relationships on shark species (Prionace glauca, Isurus oxyrinchus, Carcharhinus longimanus and Carcharhinus falciformis) were obtained from 8,331 observations recorded at sea on longliners. Significance of the sex factor was specifically assessed using GLM procedures. Linear and non-linear fits of size-weight data by species were tested. The results obtained were compared with those values provided by other authors using equivalent type of data. Deviation of the predicted versus observer weights were also assessed. Both types of fits tested have provided similar results, their confidence intervals plotted are mostly overlapped and the equations obtained were generally within those confidence intervals. Predicted mean dressed weights at size by species were in most cases quite similar or just mimetic to those obtained using equations previously reported.En prens

Measurement of the broadband complex permittivity of soils in the frequency domain with a low-cost Vector Network Analyzer and an Open-Ended coaxial probe

The performance of a handheld Vector Network Analyzer (VNA), the nanoVNA, a low-cost, open-source instrument, was evaluated. The instrument measures the complex permittivity of dielectric media from 1-port reflection parameters in the 1 – 900 MHz bandwidth. We manufactured an open-ended coaxial probe using a SMA-N coaxial adapter to perform dielectric measurements. The accuracy of the nanoVNA was comparable to that of a commercial VNA between 1 and 500 MHz according to tests in reference organic liquids, while a lack of stability was found beyond 700 MHz. The self-manufactured open-ended coaxial probe was subjected to a Finite Element Method (FEM) analysis and its electromagnetic (EM) field penetration depth was determined to be 1.5 mm at 100 MHz, being reduced to 1.3 at 900 MHz and thus demonstrating a frequency-dependent support volume. The broadband complex permittivity of three mineral soils of varied textures was obtained for a range of bulk densities and water contents from dry to water-saturated conditions. The dielectric response of the soils approximated the well-known Topp et al. (1980) equation at high frequencies. At lower frequency however, higher permittivities were exhibited due to dielectric dispersion, which emphasizes the importance of EM-based soil moisture sensor operating frequency when considering sensor calibration or comparing the response of different sensors.This research was funded by Agencia Estatal de Investigación (AEI), project numbers: AGL2016-77282-C3-3-R and PID2019-106226-C22 AEI/https:///https://doi.org/10.13039/501100011033 | Ministerio de Educación y Formación Profesional, grant numbers: FPU17/05155 and FPU19/00020. Funding for David A. Robinson was provided by a Natural Environment Research Council (NERC) award number NE/R016429/1 as part of the UK–ScaPE Programme Delivering National Capability. We also acknowledge funding from the Polish National Agency for Academic Exchange, grant number: PPI/APM/2018/1/00048/U/001. The authors wish to thank Agencia Estatal de Investigación (AEI), Ministerio de Educación y Formación Profesional, Natural Environment Research Council (NERC) and Polish National Agency for Academic Exchange (NAWA) for the funding provided. The authors also wish to thank Juan Antonio Albaladejo for his help in machining the experimental OE coaxial probe

Memory effect in triglycine sulfate induced by a transverse electric field: specific heat measurement

The influence of a transverse electric field on the specific heat of triglycine sulfate (TGS) has been studied. The specific heat of TGS has been measured on heating the sample from the ferroelectric to the paraelectric phase after prolonged application of transverse electric field (i.e. perpendicular to the ferroelectric axis). It is shown that the specific heat of TGS can 'remember' the temperature T(s) at which the transverse field was previously applied.España, Gobierno de España IS-2006-0404

Resistance to colistin and production of extended-spectrum ß-lactamases and/or AmpC enzymes in Salmonella isolates collected from healthy pigs in Northwest Spain in two periods: 2008-2009 and 2018

Salmonellosis is a common subclinical infection in pigs and therefore apparently healthy animals may represent a reservoir of antibiotic-resistant Salmonella for humans. This study estimates and characterizes resistance to two classes of antimicrobials considered of the highest priority within the critically important antimicrobials for humans, i.e. colistin (CR) and 3rd generation cephalosporins (3GC), on a collection of Salmonella isolates from pigs from two periods: between 2008 and 09, when colistin was massively used; and in 2018, after three years under a National Plan against Antibiotic Resistance. Prevalence of CR was low (6 out of 625; 0.96%; 95%CI: 0.44–2.1) in 2008–09 and associated mostly to the mcr-1 gene, which was detected in four S. 4,5,12:i:- isolates. Polymorphisms in the pmrAB genes were detected in a S. 9,12:-:- isolate. No CR was detected in 2018 out of 59 isolates tested. Among 270 Salmonella isolates considered for the assessment of resistance to 3GC in the 2008–2009 sampling, only one Salmonella Bredeney (0.37%; 95%CI: 0.07–2.1) showed resistance to 3GC, which was associated with the blaCMY-2 gene (AmpC producer). In 2018, six isolates out of 59 (10.2%; 95%CI: 4.7–20.5) showed resistance to 3GC, but only two different strains were identified (S. 4,12:i:- and S. Rissen), both confirmed as extended-spectrum β-lactamases (ESBL) producers. The blaCTX-M-3 and blaTEM-1b genes in S. 4,12:i:- and the blaTEM-1b gene in S. Rissen seemed to be associated with this resistance. Overall, the prevalence of CR in Salmonella appeared to be very low in 2008–2009 despite the considerable use of colistin in pigs at that time, and seemed to remain so in 2018. Resistance to 3GC was even lower in 2008–2009 but somewhat higher in 2018. Resistance was mostly coded by genes associated with mobile genetic elements. Most serotypes involved in these antimicrobial resistances displayed a multidrug resistance pattern and were considered zoonotic

Time Course and Magnitude of Tolerance to the Ergogenic Effect of Caffeine on the Second Ventilatory Threshold

Pre-exercise caffeine ingestion has been shown to increase the workload at ventilatory threshold, suggesting an ergogenic effect of this stimulant on submaximal aerobic exercise. However, the time course of tolerance to the effect of caffeine on ventilatory threshold is unknown. This study aimed to determine the evolution of tolerance to the ergogenic effect of caffeine on the ventilatory threshold. Methods: Eleven participants (age 32.3 ± 4.9 yrs, height 171 ± 8 cm, body mass 66.6 ± 13.6 kg, VO2max = 48.0 ± 3.8 mL/kg/min) took part in a longitudinal, double-blind, placebo-controlled, randomized, crossover experimental design. Each participant took part in two identical treatments: in one treatment, participants ingested a capsule containing 3 mg of caffeine per kg of body mass per day (mg/kg/day) for twenty consecutive days; in the other treatment, participants ingested a capsule filled with a placebo for the same duration and frequency. During these treatments, participants performed a maximal ramp test on a cycle ergometer three times per week and the second ventilatory threshold (VT2) was assessed by using the ventilatory equivalents for oxygen and carbon dioxide. Results: A two-way ANOVA with repeated measures (substance × time) revealed statistically significant main effects of caffeine (p < 0.01) and time (p = 0.04) on the wattage obtained at VT2, although there was no interaction (p = 0.09). In comparison to the placebo, caffeine increased the workload at VT2 on days 1, 4, 6 and 15 of ingestion (p < 0.05). The size of the ergogenic effect of caffeine over the placebo on the workload at VT2 was progressively reduced with the duration of the treatment. In addition, there were main effects of caffeine (p = 0.03) and time (p = 0.16) on VO2 obtained at VT2, with no interaction (p = 0.49). Specifically, caffeine increased oxygen uptake at VT2 on days 1 and 4 (p < 0.05), with no other caffeine–placebo differences afterwards. For heart rate obtained at VT2, there was a main effect of substance (p < 0.01), while the overall effect of time (p = 0.13) and the interaction (p = 0.22) did not reach statistical significance. Heart rate at VT2 was higher with caffeine than with the placebo on days 1 and 4 (p < 0.05). The size of the effect of caffeine on VO2 and heart at VT2 tended to decline over time. Conclusion: Pre-exercise intake of 3 mg/kg/day of caffeine for twenty days enhanced the wattage obtained at VT2 during cycling ramp tests for ~15 days of ingestion, while there was a progressive attenuation of the size of the ergogenic effect of caffeine on this performance variable. Therefore, habituation to caffeine through daily ingestion may reduce the ergogenic effect of this stimulant on aerobic exercise of submaximal intensity

Preclinical Characterization of Antioxidant Quinolyl Nitrone QN23 as a New Candidate for the Treatment of Ischemic Stroke

Nitrones are encouraging drug candidates for the treatment of oxidative stress-driven diseases such as acute ischemic stroke (AIS). In a previous study, we found a promising quinolylnitrone, QN23, which exerted a neuroprotective effect in neuronal cell cultures subjected to oxygen–glucose deprivation and in experimental models of cerebral ischemia. In this paper, we update the biological and pharmacological characterization of QN23. We describe the suitability of intravenous administration of QN23 to induce neuroprotection in transitory four-vessel occlusion (4VO) and middle cerebral artery occlusion (tMCAO) experimental models of brain ischemia by assessing neuronal death, apoptosis induction, and infarct area, as well as neurofunctional outcomes. QN23 significantly decreased the neuronal death and apoptosis induced by the ischemic episode in a dose-dependent manner and showed a therapeutic effect when administered up to 3 h after post-ischemic reperfusion onset, effects that remained 11 weeks after the ischemic episode. In addition, QN23 significantly reduced infarct volume, thus recovering the motor function in a tMCAO model. Remarkably, we assessed the antioxidant activity of QN23 in vivo using dihydroethidium as a molecular probe for radical species. Finally, we describe QN23 pharmacokinetic parameters. All these results pointing to QN23 as an interesting and promising preclinical candidate for the treatment of AIS.This work was supported by the Instituto de Salud Carlos III and co-financed by the European Development Regional Fund (FEDER) through grants PI18/00255, RD16/0019/0006, and RD21/0006/0019 to J.M. and A.A., and RD16/0019/0008 and RD21/0006/0014 to J.B.S.; the MINECO grant SAF2015-65586-R to J.M.-C.; and the Comunidad de Madrid Neurocentro project B2017/BMD-3760 to D.G.-N

Genes associated with metabolic syndrome predict disease-free survival in stage II colorectal cancer patients. A novel link between metabolic dysregulation and colorectal cancer

Producción CientíficaStudies have recently suggested that metabolic syndrome and its components increase the risk of colorectal cancer. Both diseases are increasing in most countries, and the genetic association between them has not been fully elucidated. The objective of this study was to assess the association between genetic risk factors of metabolic syndrome or related conditions (obesity, hyperlipidaemia, diabetes mellitus type 2) and clinical outcome in stage II colorectal cancer patients. Expression levels of several genes related to metabolic syndrome and associated alterations were analysed by real-time qPCR in two equivalent but independent sets of stage II colorectal cancer patients. Using logistic regression models and cross-validation analysis with all tumour samples, we developed a metabolic syndrome-related gene expression profile to predict clinical outcome in stage II colorectal cancer patients. The results showed that a gene expression profile constituted by genes previously related to metabolic syndrome was significantly associated with clinical outcome of stage II colorectal cancer patients. This metabolic profile was able to identify patients with a low risk and high risk of relapse. Its predictive value was validated using an independent set of stage II colorectal cancer patients. The identification of a set of genes related to metabolic syndrome that predict survival in intermediate-stage colorectal cancer patients allows delineation of a high-risk group that may benefit from adjuvant therapy and avoid the toxic and unnecessary chemotherapy in patients classified as low risk. Our results also confirm the linkage between.Ministerio de Ciencia, Innovación y Universidades (AGL2010-21565, RyC 2008-03734, IPT-2011-1248-060000),y la Comunidad de Madrid (ALIBIRD, S2009/AGR-1469

Strain effect on electronic transport and ferromagnetic transition temperature in La0.9_{0.9}Sr0.1_{0.1}MnO3_{3} thin films

We report on a systematic study of strain effects on the transport properties and the ferromagnetic transition temperature $T_{c}$ of high-quality La$_{0.9}$Sr$_{0.1}$MnO$_{3}$ thin films epitaxially grown on (100) SrTiO$_{3}$ substrates. Both the magnetization and the resistivity are critically dependent on the film thickness. $T_{c}$ is enhanced with decreasing the film thickness due to the compressive stain produced by lattice mismatch. The resistivity above 165 K of the films with various thicknesses is consistent with small polaronic hopping conductivity. The polaronic formation energy $E_{P}$ is reduced with the decrease of film thickness. We found that the strain dependence of $T_{c}$ mainly results from the strain-induced electron-phonon coupling. The strain effect on $E_{P}$ is in good agreement with the theoretical predictions.Comment: 6 pages and 5 figures, accepted for publication in Phys. Rev.

Measurement of the broadband complex permittivity of soils in the frequency domain with a low-cost vector network analyzer and an open-ended coaxial probe

The performance of a handheld Vector Network Analyzer (VNA), the nanoVNA, a low-cost, open-source instrument, was evaluated. The instrument measures the complex permittivity of dielectric media from 1-port reflection parameters in the 1 – 900 MHz bandwidth. We manufactured an open-ended coaxial probe using a SMA-N coaxial adapter to perform dielectric measurements. The accuracy of the nanoVNA was comparable to that of a commercial VNA between 1 and 500 MHz according to tests in reference organic liquids, while a lack of stability was found beyond 700 MHz. The self-manufactured open-ended coaxial probe was subjected to a Finite Element Method (FEM) analysis and its electromagnetic (EM) field penetration depth was determined to be 1.5 mm at 100 MHz, being reduced to 1.3 at 900 MHz and thus demonstrating a frequency-dependent support volume. The broadband complex permittivity of three mineral soils of varied textures was obtained for a range of bulk densities and water contents from dry to water-saturated conditions. The dielectric response of the soils approximated the well-known Topp et al. (1980) equation at high frequencies. At lower frequency however, higher permittivities were exhibited due to dielectric dispersion, which emphasizes the importance of EM-based soil moisture sensor operating frequency when considering sensor calibration or comparing the response of different sensors