Search

Phenotypic heterogeneity with variable severity has been reported in female carriers of retinitis pigmentosa GTPase regulator (RPGR) mutations, including a male-type phenotype. A phenomenon not fully understood is peripapillary retinal nerve fiber layer (pRNFL) thickening in male patients with RPGR-associated X-linked retinitis pigmentosa, especially in the temporal sector. We aim to describe the genetic spectrum, retinal phenotypes, and pRNFL thickness in a cohort of Caucasian RPGR-mutation heterozygotes.A cross-sectional study was conducted at an inherited retinal degeneration (IRD) reference center in Portugal. Female patients heterozygous for clinically significant RPGR variants were identified using the IRD-PT registry. A complete ophthalmologic examination was performed, complemented by macular and peripapillary spectral domain optical coherence tomography (SD-OCT), ultra-widefield color fundus photography (UW-CFP), and ultra-widefield fundus autofluorescence (UW-FAF). The retinal phenotypes were graded according to previously described classifications. The pRNFL thickness across the superior, inferior, nasal, and temporal quadrants was compared to the Spectralis® RNFL age-adjusted reference database.Forty-eight eyes from 24 females (10 families) were included in the study. Genetic analysis yielded 8 distinct clinically significant frameshift variants in RPGR gene, 3 of which herein reported for the first time. No association was found between mutation location and best-corrected visual acuity (BCVA) or retinal phenotype. Age was associated with worse BCVA and more advanced phenotypes on SD-OCT, UW-CFP, and UW-FAF. Seven women (29.17%) presented a male-type phenotype on UW-FAF in at least one eye. An association was found between UW-FAF and pRNFL thickness in the temporal sector (p = 0.003), with the most advanced fundus autofluorescence phenotypes showing increased pRNFL thickness in this sector.This study expands the genetic landscape of RPGR-associated disease by reporting 3 novel clinically significant variants. We have shown that clinically severe phenotypes are not uncommon among female carriers. Furthermore, we provide novel insights into pRNFL changes observed in RPGR heterozygotes that mimic what has been reported in male patients.

Background: Faecalibacterium prausnitzii, one of the most important bacteria of the human gut microbiota, produces butyrate (a short-chain fatty acid). Short-chain fatty acids are known to influence thyroid physiology and thyroid cancer’s response to treatment. We aimed to analyze the relative abundance of Faecalibacterium prausnitzii on the gut microbiota of differentiated thyroid cancer patients compared to controls and its variation after radioiodine therapy (RAIT). Methods: Fecal samples were collected from 37 patients diagnosed with differentiated thyroid cancer before and after radioiodine therapy and from 10 volunteers. The abundance of F. prausnitzii was determined using shotgun metagenomics. Results: Our study found that the relative abundance of F. prausnitzii is significantly reduced in thyroid cancer patients compared to volunteers. We also found that there was a mixed response to RAIT, with an increase in the relative and absolute abundances of this bacterium in most patients. Conclusions: Our study confirms that thyroid cancer patients present a dysbiotic gut microbiota, with a reduction in F. prausnitzii’s relative abundance. In our study, radioiodine did not negatively affect F. prausnitzii, quite the opposite, suggesting that this bacterium might play a role in resolving radiation aggression issues.

Mutations linked to neurodevelopmental disorders, such as intellectual disability (ID), are frequently found in genes that encode for proteins of the excitatory synapse. Transmembrane AMPA receptor regulatory proteins (TARPs) are AMPA receptor auxiliary proteins that regulate crucial aspects of receptor function. Here, we investigate a mutant form of the TARP family member stargazin, described in an ID patient. Molecular dynamics analyses predicted that the ID-associated stargazin variant, V143L, weakens the overall interface of the AMPAR:stargazin complex and impairs the stability of the complex. Knock-in mice harboring the V143L stargazin mutation manifest cognitive and social deficits and hippocampal synaptic transmission defects, resembling phenotypes displayed by ID patients. In the hippocampus of stargazin V143L mice, CA1 neurons show impaired spine maturation, abnormal synaptic transmission and long-term potentiation specifically in basal dendrites, and synaptic ultrastructural alterations. These data suggest a causal role for mutated stargazin in the pathogenesis of ID and unveil a new role for stargazin in the development and function of hippocampal synapses. This work was supported by a NARSAD Independent Investigator Grant (#23151) and a NARSAD Young Investigator Grant (#20733) from the Brain and Behavior Research Foundation, by a research grant from the Jérôme Lejeune Foundation (#1530), by “la Caixa” Foundation (ID 100010434), and FCT, I.P under the project code LCF/PR/HP20/52300003, by a Marie Curie Integration Grant (618525), by a Bial Foundation Grant (266/2016), by national funds through the Portuguese Science and Technology Foundation (FCT: UID/NEU/04539/2013, UIDB/04539/2020, POCI-01-0145-FEDER-28541, POCI-01-0145-FEDER-016682, PTDC/QUI-OUT/32243/2017 and CPCA/A0/7302/2020), and by the European Regional Development Fund (ERDF), through the Centro 2020 Regional Operational Programme, under project CENTRO-01-0145-FEDER-000008:BrainHealth 2020. GLC, NB, MVR, ME and CAVB were supported by FCT through Ph.D. scholarships SFRH/BD/51962/2012, SFRH/BD/144881/2019, SFRH/BD/129236/2017, SFRH/BD/51958/2012 and SFRH/BD/145457/2019, respectively. ASI and JG were supported by FCT through Postdoctoral fellowship SFRH/BPD122299/2016 and SFRH/BPD/120611/2016, respectively. RPG and RM received support from FCT/DGES, under the program “Verão com Ciência”.

Este trabalho apresenta uma revisão bibliográfica e patentária referente ao uso da narrativa visual aplicada ao ensino e à aprendizagem. A pesquisa tem caráter qualitativo e quantitativo, de forma a se obter resultados aprofundados e abrangentes. Foram utilizadas as plataformas Scopus e Web of Science para o levantamento bibliográfico. Para o levantamento patentário, a busca foi realizada nas bases Orbit®, Esp@cenet e INPI. A pesquisa mostrou que a abordagem da narrativa visual aplicada à educação vem ganhando espaço na área acadêmica, com diversos estudos que abrangem seu entendimento como ferramenta inovadora, adotando diferentes mídias e tecnologias. Essa abordagem facilita a compreensão e acessibilidade de diversos assuntos, em diferentes áreas do conhecimento, podendo ser aplicada a diferentes públicos, sendo também um possível canal de inclusão de indivíduos com diferentes tipos de limitações.

This research was funded by the Fundação para a Ciência e Tecnologia, I.P. (FCT), through project grants to S.R.P. (PTDC/BIA-PRO/109796/2009 and PTDC/BIA-BQM/29442/2017) and A.L.C. (RECI/BBB-BEP/0124/2012), and scholarship to C.S.N. (SFRH/BD/87878/2012). This work was also supported by national funds from the FCT within the scope of the project UIDP/04378/2020 and UIDB/04378/2020 of the Research Unit on Applied Molecular Biosciences—UCIBIO and the project LA/P/0140/2020 of the Associate Laboratory Institute for Health and Bioeconomy—i4HB. Acknowledgments The authors would like to thank Lina Juknaité for her contribution in the initial crystallization studies. The authors acknowledge the European Synchrotron Radiation Facility and the Swiss Light Source for provision of synchrotron radiation facilities and access to beamlines BM30 and PXIII (X06DA), respectively. Neisseria gonorrhoeae is an obligate human pathogenic bacterium responsible for gonorrhea, a sexually transmitted disease. The bacterial peroxidase, an enzyme present in the periplasm of this bacterium, detoxifies the cells against hydrogen peroxide and constitutes one of the primary defenses against exogenous and endogenous oxidative stress in this organism. The 38 kDa heterologously produced bacterial peroxidase was crystallized in the mixed-valence state, the active state, at pH 6.0, and the crystals were soaked with azide, producing the first azide-inhibited structure of this family of enzymes. The enzyme binds exogenous ligands such as cyanide and azide, which also inhibit the catalytic activity by coordinating the P heme iron, the active site, and competing with its substrate, hydrogen peroxide. The inhibition constants were estimated to be 0.4 ± 0.1 µM and 41 ± 5 mM for cyanide and azide, respectively. Imidazole also binds and inhibits the enzyme in a more complex mechanism by binding to P and E hemes, which changes the reduction potential of the latest heme. Based on the structures now reported, the catalytic cycle of bacterial peroxidases is revisited. The inhibition studies and the crystal structure of the inhibited enzyme comprise the first platform to search and develop inhibitors that target this enzyme as a possible new strategy against N. gonorrhoeae. publishersversion published

Proteasome activity at the excitatory synapse plays an important role in neuronal communication. The proteasome translocation to synapses is mediated by neuronal activity, in particular the activation of N-methyl-d-aspartate receptors (NMDARs). These receptors are composed of different subunits with distinct trafficking properties that provide various signalling and plasticity features to the synapse. Yet whether the interplay between the proteasome and NMDAR relies on specific subunit properties remain unclear. Using a combination of single molecule and immunocytochemistry imaging approaches in rat hippocampal neurons, we unveil a specific interplay between GluN2B-containing NMDARs (GluN2B-NMDARs) and the synaptic proteasome. Sustained proteasome activation specifically increases GluN2B-NMDAR (not GluN2A-NMDAR) lateral diffusion. In addition, when GluN2B-NMDAR expression is downregulated, the proteasome localization decreases at glutamatergic synapses. Collectively, our data fuel a model in which the cellular dynamics and location of GluN2B-NMDARs and proteasome are intermingled, shedding new lights on the NMDAR-dependent regulation of synaptic adaptation.

the Associate Laboratory for Animal and Veterinary Sciences (AL4AnimalS) grant LA/P/0059/2020. LA/P/0140/2020 of the Associate Laboratory Institute for Health and Bioeconomy - i4HB. National Institutes of Health R01-GM129325 and the Office of Cyber Infrastructure and Computational Biology, National Institute of Allergy and Infectious Diseases. Publisher Copyright: © 2022 The Author(s) The cellulosome is an elaborate multi-enzyme structure secreted by many anaerobic microorganisms for the efficient degradation of lignocellulosic substrates. It is composed of multiple catalytic and non-catalytic components that are assembled through high-affinity protein-protein interactions between the enzyme-borne dockerin (Doc) modules and the repeated cohesin (Coh) modules present in primary scaffoldins. In some cellulosomes, primary scaffoldins can interact with adaptor and cell-anchoring scaffoldins to create structures of increasing complexity. The cellulosomal system of the ruminal bacterium, Ruminococcus flavefaciens, is one of the most intricate described to date. An unprecedent number of different Doc specificities results in an elaborate architecture, assembled exclusively through single-binding-mode type-III Coh-Doc interactions. However, a set of type-III Docs exhibits certain features associated with the classic dual-binding mode Coh-Doc interaction. Here, the structure of the adaptor scaffoldin-borne ScaH Doc in complex with the Coh from anchoring scaffoldin ScaE is described. This complex, unlike previously described type-III interactions in R. flavefaciens, was found to interact in a dual-binding mode. The key residues determining Coh recognition were also identified. This information was used to perform structure-informed protein engineering to change the electrostatic profile of the binding surface and to improve the affinity between the two modules. The results show that the nature of the residues in the ligand-binding surface plays a major role in Coh recognition and that Coh-Doc affinity can be manipulated through rational design, a key feature for the creation of designer cellulosomes or other affinity-based technologies using tailored Coh-Doc interactions. publishersversion published

PD/BD/105753/2014 PTDC/BII-BIO/28878/2017 UIDB/04378/ 2020 POCI-01-0145-FEDER-007728 After more than one hundred and thirty thousand protein structures determined by X-ray crystallography, the challenge of protein crystallization for 3D structure determination remains. In the quest for additives for efficient protein crystallization, inorganic materials emerge as an alternative. Magnetic particles (MPs) are versatile inorganic materials, easy to use, modify and manipulate in a wide range of biological assays. The potential of using functionalised MPs as crystallization chaperones for protein crystallization was shown in this work. MPs with distinct coatings were rationally designed to promote protein crystallization by affinity-triggered heterogeneous nucleation. Hen egg white lysozyme (HEWL) and trypsin, were crystallized in the presence of MPs either bare or coated with a polysaccharide (chitin) or a protein (casein), respectively. The addition of MPs was characterized in terms of bound protein to the MPs, crystal morphology, time-lapse of crystal emergence, crystallization yield fold change and crystal diffraction quality for structure determination. The MPs additives have shown to bind to the respective target protein, and to promote nucleation and crystal growth without compromising crystal morphology. On the other hand, MPs addition led to faster detectable crystal emergence and up to 13 times higher crystallization yield, addressing some the challenges in protein crystallization, the main bottleneck of macromolecular crystallography. Structure determination of the protein crystallized in the presence of MPs revealed that the structural characteristics of the protein remained unchanged, as shown by the superposition with PDB annotated proteins. Moreover, and unlike most reported cases, it was possible to exclude the inhibitor benzamidine during trypsin crystallisation, which is a remarkable result opening new prospects in enzyme engineering and drug design. Our results show that MPs coated with affinity ligands to target proteins can be used as controlled and tailor-made crystallization inducers. authorsversion published

Objectives The involvement of adolescents in violent behaviors is becoming an important public health problem that is concerning physicians as it is associated with a decrease in adult life quality. In this context, bullying is a type of aggressive behavior that occurs repeatedly in interpersonal relationship where power imbalance exists increasing the risk of physical and psychosocial problems in all its intervenient. Prevalence of bullying in Portuguese adolescents is estimated to be ranging from 16.2 to 27.5%. Sleep disorders are a well-known problem related to bullying situations, either as a consequence or a risk factor; estimated prevalence in adolescence is 25% worldwide. In the present study we aimed to investigate the association between sleep quality and the involvement in school bullying in Portuguese adolescents in two high schools. Methods Data collected from a cross-sectional survey addressing questions on bullying behaviors and sleep quality was analyzed using SPSS 22.0. A two-tailed p-value of less than 0.05 was considered significant for all tests. Results We have included 171 adolescents, mean age of 14.39 ± 1.149 years, 59.6% were boys; prevalence of bullying behaviors was 28.1%. Among the students 15.2% were identified as “pure victims”, 8.8% as “pure bullies”, 4.1% as “bully-victims” and 71.9% as “neutrals”. “Pure bullies” and “bully-victims” present higher Athens Insomina Scale (AIS) scores (p = 0.004) and higher prevalence of insomnia (p = 0.004). The prevalence of insomnia was significantly higher in the group involved in bullying behaviors comparing with the neutrals (58.3% vs 30%, p = 0.001). We verified that 65.4% of the reported bullying situations took place inside the school and 8.8% of those teachers did nothing to help the victim and in 17.5% school staff took the same attitude. Conclusions We found a prevalence of bullying behaviors higher than the one reported in previous studies and a prevalence of insomnia of 38.0%, proving that bullying and sleep disturbances are two major problems in adolescence which may increase future risk for psychiatric symptoms, involvement in criminal activities, hyperactive, and inattentive behaviors. We verified a strong relation between insomnia and involvement in bullying situations with those who are involved reporting difficulties in sleep induction. The group of “bully-victims” was identified as potentially being the one with the most troubled adolescents since they combine characteristics of both bullies and victims. The authors conclude that there should be an investment in school-based interventions that help recognize and reduce school bullying and change lifestyle behaviors that may negatively influence adolescents' development.

No. SCENT‐ERC‐2014‐STG‐639123, 2015–2022) UIDP/04378/2020 UIDB/04378/2020 LA/P/0140/2020 SFRH/BD/113112/2015 Relative humidity is simultaneously a sensing target and a contaminant in gas and volatile organic compound (VOC) sensing systems, where strategies to control humidity interference are required. An unmet challenge is the creation of gas-sensitive materials where the response to humidity is controlled by the material itself. Here, humidity effects are controlled through the design of gelatin formulations in ionic liquids without and with liquid crystals as electrical and optical sensors, respectively. In this design, the anions [DCA]− and [Cl]− of room temperature ionic liquids from the 1-butyl-3-methylimidazolium family tailor the response to humidity and, subsequently, sensing of VOCs in dry and humid conditions. Due to the combined effect of the materials formulations and sensing mechanisms, changing the anion from [DCA]− to the much more hygroscopic [Cl]−, leads to stronger electrical responses and much weaker optical responses to humidity. Thus, either humidity sensors or humidity-tolerant VOC sensors that do not require sample preconditioning or signal processing to correct humidity impact are obtained. With the wide spread of 3D- and 4D-printing and intelligent devices, the monitoring and tuning of humidity in sustainable biobased materials offers excellent opportunities in e-nose sensing arrays and wearable devices compatible with operation at room conditions. publishersversion published

This work presents a bibliographic and patent review regarding the use of visual narrative applied to teaching and learning. The research is qualitative and quantitative, aiming to obtain in-depth and comprehensive results. Scopus and Web of Science platforms were used for the bibliographic research. For the patent review, the search was carried out on the Orbit®, Esp@cenet and INPI databases. The research showed that the visual narrative approach, applied to education, has been gaining ground in the academic field, with several studies that cover its understanding as an innovative tool, adopting different media and technologies. This approach facilitates the understanding and accessibility of various subjects, in different areas of knowledge, and can be applied to different audiences, being also a possible channel for the inclusion of individuals with different types of limitations. Este trabalho apresenta uma revisão bibliográfica e patentária referente ao uso da narrativa visual aplicada ao ensino e à aprendizagem. A pesquisa tem caráter qualitativo e quantitativo, de forma a se obter resultados aprofundados e abrangentes. Foram utilizadas as plataformas Scopus e Web of Science para o levantamento bibliográfico. Para o levantamento patentário, a busca foi realizada nas bases Orbit®, Esp@cenet e INPI. A pesquisa mostrou que a abordagem da narrativa visual aplicada à educação vem ganhando espaço na área acadêmica, com diversos estudos que abrangem seu entendimento como ferramenta inovadora, adotando diferentes mídias e tecnologias. Essa abordagem facilita a compreensão e acessibilidade de diversos assuntos, em diferentes áreas do conhecimento, podendo ser aplicada a diferentes públicos, sendo também um possível canal de inclusão de indivíduos com diferentes tipos de limitações.

CNGB1 gene mutations are a well-known cause of autosomal recessive retinitis pigmentosa (RP), which was recently associated with olfactory dysfunction. The purpose of this study was to report the molecular spectrum and the ocular and olfactory phenotypes of a multiethnic cohort with CNGB1-associated RP. A cross-sectional case series was conducted at two ophthalmic genetics referral centers. Consecutive patients with molecularly confirmed CNGB1-related RP were included. All patients underwent a complete ophthalmological examination complemented by psychophysical olfactory evaluation. Fifteen patients (10 families: 8 Portuguese, 1 French, and 1 Turkish), mean aged 57.13 ± 15.37 years old (yo), were enrolled. Seven disease-causing variants were identified, two of which are reported for the first time: c.2565_2566del and c.2285G > T. Although 11/15 patients reported onset of nyctalopia before age 10, diagnosis was only established after 30 yo in 9/15. Despite widespread retinal degeneration being present in 14/15 probands, a relatively preserved visual acuity was observed throughout follow-up. Olfactory function was preserved in only 4/15 patients, all of whom carried at least one missense variant. Our study supports previous reports of an autosomal recessive RP-olfactory dysfunction syndrome in association with certain disease-causing variants in the CNGB1 gene and expands the mutational spectrum of CNGB1-related disease by reporting two novel variants.

International audience; The SOUL, or heme-binding protein HBP/SOUL, family represents a group of evolutionary conservedputative heme-binding proteins that contains a number of members in animal, plant and bacterial spe-cies. The structures of the murine form of HEBP1, or p22HBP, and the human form of HEBP2, or SOUL,have been determined in 2006 and 2011 respectively.In this work we discuss the structures of HEBP1 and HEBP2 in light of new X-ray data for heme boundmurine HEBP1. The interaction between tetrapyrroles and HEBP1, initially proven to be hydrophobic innature, was thought to also involve electrostatic interactions between heme propionate groups and pos-itively charged amino acid side chains. However, the new X-ray structure, and results from murine HEBP1variants and human HEBP1, confirm the hydrophobic nature of the heme-HEBP1 interaction, resulting inKdvalues in the low nanomolar range, and rules out any electrostatic stabilization. Results from NMRrelaxation time measurements for human HEBP1 describe a rigid globular protein with no change inmotional regime upon heme binding.X-ray structures deposited in the PDB for human HEBP2 are very similar to each other and to the newheme-bound murine HEBP1 X-ray structure (backbone rmsd ca. 1 Å). Results from a HSQC spectrum cen-tred on the histidine side chain Nd-proton region for HEBP2 confirm that HEBP2 does not bind heme viaH42 as no chemical shift differences were observed upon heme addition for backbone NH and Ndprotons.A survey of the functions attributed to HEBP1 and HEBP2 over the last 20 years span a wide range ofcellular pathways. Interestingly, many of them are specific to higher eukaryotes, particularly mammalsand a potential link between heme release under oxidative stress and human HEBP1 is also examinedusing recent data. However, at the present moment, trying to relate function to the involvement of hemehttps://doi.org/10.1016/j.ccr.2021.2141890010-8545/Ó2021 Elsevier B.V. All rights reserved.Abbreviations:HEBP1, Heme-binding protein 1 (or p22HBP); HEBP2, Heme-binding protein 2 (or SOUL); BH3, Bcl-2 homology 3; HSQC, heteronuclear single quantumcoherence spectroscopy; TROSY, transverse relaxation optimized spectroscopy; FQ, fluorescence quenching; PPIX, protoporphyrin IX; sGC, solubleGuanylyl Cyclase; hetNOE,heteronuclear nuclear Overhauser effect; rmsd, root mean squared deviation; BMRB, biological magnetic resonance bank; VAST, vector alignment search tool; MEL, murineerythroleukemia.⇑Corresponding authors.E-mail addresses:brian.goodfellow@ua.pt(B.J. Goodfellow),mdam@fct.unl(A.L. Macedo).Coordination Chemistry Reviews 448 (2021) 214189Contents lists available atScienceDirectCoordination Chemistry Reviewsjournal homepage: www.elsevier.com/locate/ccr

contract DL-57/2016 WT108430/Z/15/Z WT218304/Z/19/Z 22-FY18-821 A multigene polysaccharide utilization locus (PUL) encoding enzymes and surface carbohydrate (glycan)-binding proteins (SGBPs) was recently identified in prominent members of Bacteroidetes in the human gut and characterized in Bacteroides ovatus. This PUL-encoded system specifically targets mixed-linkage β1,3-1,4-glucans, a group of diet-derived carbohydrates that promote a healthy microbiota and have potential as prebiotics. The BoSGBPMLG-A protein encoded by the BACOVA_2743 gene is a SusD-like protein that plays a key role in the PUL's specificity and functionality. Here, we perform a detailed analysis of the molecular determinants underlying carbohydrate binding by BoSGBPMLG-A, combining carbohydrate microarray technology with quantitative affinity studies and a high-resolution X-ray crystallography structure of the complex of BoSGBPMLG-A with a β1,3-1,4-nonasaccharide. We demonstrate its unique binding specificity toward β1,3-1,4-gluco-oligosaccharides, with increasing binding affinities up to the octasaccharide and dependency on the number and position of β1,3 linkages. The interaction is defined by a 41-Å-long extended binding site that accommodates the oligosaccharide in a mode distinct from that of previously described bacterial β1,3-1,4-glucan-binding proteins. In addition to the shape complementarity mediated by CH-π interactions, a complex hydrogen bonding network complemented by a high number of key ordered water molecules establishes additional specific interactions with the oligosaccharide. These support the twisted conformation of the β-glucan backbone imposed by the β1,3 linkages and explain the dependency on the oligosaccharide chain length. We propose that the specificity of the PUL conferred by BoSGBPMLG-A to import long β1,3-1,4-glucan oligosaccharides to the bacterial periplasm allows Bacteroidetes to outcompete bacteria that lack this PUL for utilization of β1,3-1,4-glucans. IMPORTANCE With the knowledge of bacterial gene systems encoding proteins that target dietary carbohydrates as a source of nutrients and their importance for human health, major efforts are being made to understand carbohydrate recognition by various commensal bacteria. Here, we describe an integrative strategy that combines carbohydrate microarray technology with structural studies to further elucidate the molecular determinants of carbohydrate recognition by BoSGBPMLG-A, a key protein expressed at the surface of Bacteroides ovatus for utilization of mixed-linkage β1,3-1,4-glucans. We have mapped at high resolution interactions that occur at the binding site of BoSGBPMLG-A and provide evidence for the role of key water-mediated interactions for fine specificity and affinity. Understanding at the molecular level how commensal bacteria, such as prominent members of Bacteroidetes, can differentially utilize dietary carbohydrates with potential prebiotic activities will shed light on possible ways to modulate the microbiome to promote human health. publishersversion published

Retinitis pigmentosa (RP) corresponds to a group of inherited retinal disorders where progressive rod-cone degeneration is observed. Cystoid macular edema (CME) and vitreomacular interface disorders (VMID) are known to complicate the RP phenotype, challenging an age-old concept of retained central visual acuity. The reported prevalence of these changes varies greatly among different studies. We aim to describe the frequency of CME and VMID and identify predictors of these changes in a cohort of Caucasian patients with genetically solved syndromic (sRP) and non-syndromic RP (nsRP).Cross-sectional study of patients with genetically solved sRP or nsRP. Genetic testing was clinically oriented in all probands and coordinated by a medical geneticist. The presence/absence of CME and VMIDs such as epiretinal membrane (ERM), vitreomacular traction (VMT), lamellar hole (LH), macular hole (MH), and macular pseudohole (MPH), and the integrity of the neurosensory retina and retinal pigment epithelium were evaluated in individual macular SD-OCT b-scans. Mixed-effects regression analysis models were used to identify significant predictors of BCVA, CME, and VMID. Significance was considered at α 0.05.We included 250 eyes from 125 patients. Mean age was 44.9 ± 15.7 years and 55.2% were male. Eighty-eight patients had nsRP and 37 had sRP. Median BCVA was 0.5 (0.2-1.3) logMAR. CME was found in 17.1% of eyes, while ERM was found in 54.3% of eyes. The frequency of CME (p = 0.45) and ERM (p = 0.07) did not differ between sRP and nsRP patients, nor across different inheritance patterns. Mixed-effects univariate linear regression identified age (p = 0.04), cataract surgery (p 0.01), and loss of integrity of outer retinal layers (p 0.01) as significant predictors of lower visual acuity, while increased foveal thickness (p 0.01) and the presence of CME (p = 0.04) were predictors of higher visual acuity. On mixed-effects multivariable analysis, only increased foveal thickness was significantly associated with better visual acuity (p 0.01).We found that the burden of ERM and CME in RP patients is high, highlighting the importance of screening for these potentially treatable conditions to improve the quality of life of RP patients.

Introduction: The purpose of this study was to compare clinical/demographic functional testing and multimodal imaging features between genetically solved and genetically unsolved nonsyndromic retinitis pigmentosa (nsRP) patients. Methods: A cross-sectional study was conducted at an inherited retinal dystrophies reference center. Consecutive patients with nsRP and available genetic testing results performed between 2018 and 2020 were included. Genetic testing was clinically oriented, and variants were classified according to the American College of Medical Genetics and Genomics. Only class IV or V variants were considered disease-causing. Clinical/demographic, functional, and imaging features were compared between genetically unsolved (G1) and genetically solved (G2) patients. Results: A total of 175 patients (146 families) were included: 68 patients (59 families) in G1 and 107 patients (87 families) in G2. First symptoms Conclusion: Individual clinical/demographic functional testing and multimodal imaging features should be considered when counseling patients about the probability of identifying disease-causing variants.

Understanding the specific molecular interactions between proteins and β1,3-1,4-mixed-linked d-glucans is fundamental to harvest the full biological and biotechnological potential of these carbohydrates and of proteins that specifically recognize them. The family 11 carbohydrate-binding module from Clostridium thermocellum (CtCBM11) is known for its binding preference for β1,3-1,4-mixed-linked over β1,4-linked glucans. Despite the growing industrial interest of this protein for the biotransformation of lignocellulosic biomass, the molecular determinants of its ligand specificity are not well defined. In this report, a combined approach of methodologies was used to unravel, at a molecular level, the ligand recognition of CtCBM11. The analysis of the interaction by carbohydrate microarrays and NMR and the crystal structures of CtCBM11 bound to β1,3-1,4-linked glucose oligosaccharides showed that both the chain length and the position of the β1,3-linkage are important for recognition, and identified the tetrasaccharide Glcβ1,4Glcβ1,4Glcβ1,3Glc sequence as a minimum epitope required for binding. The structural data, along with site-directed mutagenesis and ITC studies, demonstrated the specificity of CtCBM11 for the twisted conformation of β1,3-1,4-mixed-linked glucans. This is mediated by a conformation-selection mechanism of the ligand in the binding cleft through CH-π stacking and a hydrogen bonding network, which is dependent not only on ligand chain length, but also on the presence of a β1,3-linkage at the reducing end and at specific positions along the β1,4-linked glucan chain. The understanding of the detailed mechanism by which CtCBM11 can distinguish between linear and mixed-linked β-glucans strengthens its exploitation for the design of new biomolecules with improved capabilities and applications in health and agriculture. DATABASE: Structural data are available in the Protein Data Bank under the accession codes 6R3M and 6R31.

Autism spectrum disorder (ASD) is characterized by dysfunction in social interactions, stereotypical behaviours and high co-morbidity with intellectual disability. A variety of syndromic and non-syndromic neurodevelopmental disorders have been connected to alterations in metabotropic glutamate receptor (mGluR) signalling. These receptors contribute to synaptic plasticity, spine maturation and circuit development. Here, we investigate the physiological role of Gprasp2, a gene linked to neurodevelopmental disabilities and involved in the postendocytic sorting of G-protein-coupled receptors. We show that Gprasp2 deletion leads to ASD-like behaviour in mice and alterations in synaptic communication. Manipulating the levels of Gprasp2 bidirectionally modulates the surface availability of mGluR5 and produces alterations in dendritic complexity, spine density and synaptic maturation. Loss of Gprasp2 leads to enhanced hippocampal long-term depression, consistent with facilitated mGluR-dependent activation. These findings demonstrate a role for Gprasp2 in glutamatergic synapses and suggest a possible mechanism by which this gene is linked to neurodevelopmental diseases., GPRASP2 plays a role in trafficking of GPCRs and mutations in this gene have been linked to neurodevelopmental disorders. Here the authors study the role of Gprasp2 in the CNS and show that it regulates the surface availability of mGluR5 receptors and that knockout mice for this protein show autistic-like behavioural abnormalities.

Interactions of glycan-specific epitopes to human lectin receptors represent novel immune checkpoints for investigating cancer and infection diseases. By employing a multidisciplinary approach that combines isothermal titration calorimetry, NMR spectroscopy, molecular dynamics simulations, and X-ray crystallography, we investigated the molecular determinants that govern the recognition of the tumour and pathogenic glycobiomarker LacdiNAc (GalNAcβ1-4GlcNAc, LDN), including their comparison with the ubiquitous LacNAc epitope (Galβ1-4GlcNAc, LN), by two human immune-related lectins, galectin-3 (hGal-3) and the macrophage galactose C-type lectin (hMGL). A different mechanism of binding and interactions was observed for the hGal-3/LDN and hMGL/LDN complexes, which explains the remarkable difference in the binding specificity of LDN and LN by these two lectins. The new structural clues reported herein are fundamental for the chemical design of mimetics targeting hGal-3/hMGL recognition process.

The biological signals of hunger, satiety, and memory are interconnected. The role of the hormone ghrelin in regulating feeding and memory makes ghrelin receptors attractive targets for associated disorders. We investigated the effects of the high ligand-independent activity of the ghrelin receptor GHS-R1a on the physiology of excitatory synapses in the hippocampus. Blocking this activity produced a decrease in the synaptic content of AMPA receptors in hippocampal neurons and a reduction in GluA1 phosphorylation at Ser. Reducing the ligand-independent activity of GHS-R1a increased the surface diffusion of AMPA receptors and impaired AMPA receptor–dependent synaptic delivery induced by chemical long-term potentiation. Accordingly, we found that blocking this GHS-R1a activity impaired spatial and recognition memory in mice. These observations support a role for the ligand-independent activity of GHS-R1a in regulating AMPA receptor trafficking under basal conditions and in the context of synaptic plasticity that underlies learning., NARSAD Independent Investigator Grant from the Brain and Behavior Research Foundation, by national funds through the Portuguese Science and Technology Foundation (FCT; POCI-01-0145-FEDER-007440, POCI-01-0145-FEDER-PTDC/SAUNMC/4888/2014, POCI-01-0145-FEDER-28541, POCI-01-0145-FEDER-022122, and UIDB/04539/2020), and by the European Regional Development Fund (ERDF), through the Centro 2020 Regional Operational Programme under project CENTRO-01-0145-FEDER-000008:BrainHealth 2020

Background The development of multicenter patient registries promotes the generation of scientific knowledge by using real-world data. A country-wide, web-based registry for inherited retinal dystrophies (IRDs) empowers patients and community organizations, while supporting formal partnerships research. We aim to describe the design, development and deployment of a country-wide, with investigators and stakeholders in the global aim to develop high-value, high-utility web-based, user-friendly and interoperable registry for IRDs—the IRD-PT. Results The IRD-PT is a clinical/genetic research registry included in the retina.pt platform (https://www.retina.com.pt), which was developed by the Portuguese Retina Study Group. The retina.pt platform collects data on individuals diagnosed with retinal diseases, from several sites across Portugal, with over 1800 participants and over 30,000 consultations to date. The IRD-PT module interacts with the retina.pt core system which provides a range of basic functions for patient data management, while the IRD-PT module allows data capture for the specific purpose of IRDs. All IRDs are coded accordingly to the International Statistical Classification of Diseases and Related Health Problems (ICD) 9, ICD 10, ICD 11, and Orphanet Rare Disease Ontology (ORPHA codes) to make the IRD-PT interoperable with other IRD registries across the world. Furthermore, the genes are coded according to the Ontology of Genes and Genomes and Online Mendelian Inheritance in Man, whereas signs and symptoms are coded according to the Human Phenotype Ontology. The IRD-PT module pre-launched at Centro Hospitalar e Universitário de Coimbra, the largest reference center for IRDs in Portugal. As of April 1st 2020, finalized data from 537 participants were available for this preliminary analysis. Conclusions In the specific field of rare diseases, the use of registries increases research accessibility for individuals, while providing clinicians/investigators with a coherent data ecosystem necessary to boost research. Appropriate design and implementation of patient registries enables rapid decision making and ongoing data mining, ultimately leading to improved patient outcomes. We have described here the principles behind the design, development and deployment of a web-based, user-friendly and interoperable software tool aimed to generate important knowledge and collecting high-quality data on the epidemiology, genomic landscape and natural history of IRDs in Portugal.

The ability of animals to store and retrieve food caches in the wild requires the integration of biological signals of hunger, satiety and memory. The role of ghrelin in regulating feeding and memory makes ghrelin receptors an important target to shape the required cellular and molecular responses. We investigated the effects of the high ligand-independent activity of the ghrelin receptor on the physiology of excitatory synapses. Blocking this type of activity produced a decrease in the synaptic content of AMPA receptors in hippocampal neurons and a reduction in GluA1 phosphorylation at Ser845. Impaired constitutive activity from the ghrelin receptor increased surface diffusion of AMPA receptors and impaired AMPA receptor synaptic delivery mediated by chemical long-term potentiation. These observations support a role for the constitutive activity of the ghrelin receptor in regulating AMPA receptor trafficking under basal conditions and synaptic plasticity. Accordingly, we found that blocking the ghrelin receptor constitutive activity impairs spatial and recognition memory.Impact statementThis work uncovers a role for the constitutive activity of the ghrelin receptor in memory, and in the regulation of the synaptic levels of AMPA receptors, their mobility and synaptic plasticity. Underscoring the importance of deciphering the physiological role of constitutive ghrelin receptor activity, ghrelin receptor inverse agonism is now being considered as a therapy to treat alcohol use disorder.

Submitted by denison pereira (denison.rolim@uniceub.br) on 2021-01-19T14:06:54Z No. of bitstreams: 1 Ana Santos 21600205 (2).pdf: 273701 bytes, checksum: d468e66d07d7133dde660e35c45e8440 (MD5) Approved for entry into archive by Fernanda Weschenfelder (fernanda.weschenfelder@uniceub.br) on 2021-03-23T15:24:40Z (GMT) No. of bitstreams: 1 Ana Santos 21600205 (2).pdf: 273701 bytes, checksum: d468e66d07d7133dde660e35c45e8440 (MD5) Made available in DSpace on 2021-03-23T15:24:40Z (GMT). No. of bitstreams: 1 Ana Santos 21600205 (2).pdf: 273701 bytes, checksum: d468e66d07d7133dde660e35c45e8440 (MD5) Previous issue date: 2021-01-19 Este artigo tem como objetivo analisar qual seria o impacto da humanização por meio da inserção das boas práticas dispostas na Lei 11.340/2006 (Lei Maria da Penha) no processo penal. Trata-se de uma pesquisa teórica que teve como marco a Lei 11.340/2006, o livro Processo Penal Feminista da autora Soraia da Rosa Mendes que aborda de forma sensível a falta de preparação das redes de atendimento à mulher em situação de violência doméstica desde a delegacia até o juizado, bem como artigos digitais que fundamentaram o impacto do patriarcado na participação sociocultural da mulher e como isso reflete diretamente na violência de gênero. Conclui-se que a Lei Maria da Penha é completa em seus dispositivos, restando apenas a efetiva implementação das boas práticas para que a mulher vítima, de fato, se sinta acolhida em todos os momentos da busca por proteção do Sistema de Justiça Criminal, assim como antes e depois da violência.

The family 81 glycoside hydrolase (GH81) from Clostridium thermocellum is a β-1,3-glucanase belonging to cellulosomal complex. The gene encoding GH81 from Clostridium thermocellum (CtLam81A) was cloned and expressed displaying a molecular mass of ~82 kDa. CtLam81A showed maximum activity against laminarin (100 U/mg), followed by curdlan (65 U/mg), at pH 7.0 and 75 °C. CtLam81A displayed Km, 2.1 ± 0.12 mg/ml and Vmax, 109 ± 1.8 U/mg, against laminarin under optimized conditions. CtLam81A activity was significantly enhanced by Ca2+ or Mg2+ ions. Melting curve analysis of CtLam81A showed an increase in melting temperature from 91 °C to 96 °C by Ca2+ or Mg2+ ions and decreased to 82 °C by EDTA, indicating that Ca2+ and Mg2+ ions may be involved in catalysis and in maintaining structural integrity. TLC and MALDI-TOF analysis of β-1,3-glucan hydrolysed products released initially, showed β-1,3-glucan-oligosaccharides degree of polymerization (DP) from DP2 to DP7, confirming an endo-mode of action. The catalytically inactive mutant CtLam81A-E515A generated by site-directed mutagenesis was co-crystallized and tetragonal crystals diffracting up to 1.4 A resolution were obtained. CtLam81A-E515A contained 15 α-helices and 38 β-strands forming a four-domain structure viz. a β-sandwich domain I at N-terminal, an α/β-domain II, an (α/α)6 barrel domain III, and a small 5-stranded β-sandwich domain IV.

Synaptic scaling allows neurons to adjust synaptic strength in response to chronic alterations in neuronal activity. A new study in PLOS Biology identifies a pathway that synergizes protein synthesis and degradation with remodeling of the microRNA (miRNA)-induced silencing complex (miRISC) to mediate synaptic scaling.