Structure of monomeric transthyretin carrying the clinically important T119M mutation.

Mutations in the protein transthyretin can cause as well as protect individuals from transthyretin amyloidosis, an incurable fatal inherited disease. Little is known, however, about the structural basis of pathogenic and clinically protective transthyretin mutants. Here we determined the solution structure of a transthyretin monomer that carries the clinically important T119M mutation. The structure displays a non-native arrangement that is distinct from all known structures of transthyretin and highlights the importance of high-resolution studies in solution for understanding molecular processes that lead to amyloid diseases

Extraction and spectrophotometric and fluorimetric determination of micrograms of zinc in plants

9 pág.[ES] Se han aplicado dos nuevos métodos originales, espectrofotométrico y fluorimétrico, a la determinación de microgramos de cinc en material vegetal. Después de la disolución de la muestra, el cinc se extrae en solución 0,05 M de trioctilamina en tolueno, desde soluciones 1-3 M HCl en fase acuosa. El desarrollo del color o de la fluorescencia se efectua en la misma fase orgánica de la extracción sin efectuar re-extracción a fase acuosa, mediante adición de solución de oxina en N,N-dimetilformamida (DMF) y de solución de t-butil-amina en DMF. La absorbancia se mide a 403 nm. La ley de Beer se cumple en el intervalo de 1 a 10 mg. ml-1 de cinc en la fase orgánica final. La desviación estandar relativa para diez determinaciones de 50 mg. de cinc fue de 0,8%. En el método fluorimétrico, se ha empleado una radiación de excitación de 403 nm de longitud de onda y una emisión de fluorescencia de 450 nm. La intensidad relativa de fluorescencia de 450 nm. La intensidad relativa de fluorescencia era lineal con la concentración de cinc en el intervalo de 0,01 a 1 mg. ml-1) en la fase orgánica final. La desviación estandar relativa en la determinación fluorimétrica para diez determinaciones de 0,5 mg. de cinc fué de 2,2%. El valor medio de la recuperación de 0,5 mg de cinc añadidos a las soluciones de las muestras fué del 98,5%. Los resultados obtenidos en la determinación de cinc por ambos métodos propuestos en muestras de material vegetal suministradas amablemente por M. PINTA del Comité Inter-Institutos de Análisis Foliar, muestran buena precisión y exactitud.[EN] Two new spectrophotometric and fluorimetric methods for micrograms determination of zinc have been applied to plants analysis. After dissolution of the sample, zinc was extracted into 0.05 M trioctylamine in toluene solution from 1-3 M HCl aqueous solution. The development of the colour or the fluorescence was carried out in the same organic phase of the extraction, without back-extraction into aqueous phase, by addition of oxine solution in N, N-dimenthylformamide and t-butylamine in DMF solution. In the spectrophotometric method the absorbance was measured at 403 nm. Beer's law was obeyed in the range from 1 up to 10µg. ml-1 of zinc in the organic phase. The relative standard deviation for ten determinations of 50 µg. of zinc was 0,8%. In the f1uorimetric method a wavelength excitation radiation of 403 nm and a wavelength fluorescence emission of 450 nm were used. The relative intensity of f1uorescence was linear with the concentration of zinc in the range from 0.01 up to 1 µg. ml-1 of zinc in the organic phase. The standard relative deviation in the f1uorimetric determination for ten determinations of 0.5 µg of zinc was 2.2%. The mean recovery of 0.5 µg of zinc in spiked solutions was 98.5%. The results obtained for the determinations of zinc by both proposed methods in plants supplied by M. PINTA (Comité Inter-Instituts pour l'Analyse Foliaire) show good accuracy and precision.Peer reviewe

Quandoque bonus dormitat Corominas: Sobre arragua, laude, matar

Caro Baroja suggested that Basque arragua ‘melting pot’ goes back etymologically to arrugia ‘gallery in a mine’, a word mentioned by Pliny. This totally aberrant etymology is mentioned by Coromines in both the DCELC and the DCECEH. For laude ‘burial stone’, he follows Covarrubias in accepting laudem ‘praise’, but the more plausible etymology is lapidem ‘stone’. However, when it comes to the etymology of matar ‘to kill’, Coromines rejects Covarrubias’ proposal mactare on behalf of implausible phonetic change; however, mactare was a very popular word, and, despite Coromines’ rejection, has to be considered as the best candidate for an etymology for matar.Keywords: Coromines / Joan, etymology, lexicography, Spanish language, Catalan language, Latin language, Basque language

Crowdsourcing malaria parasite quantification: an online game for analyzing images of infected thick blood smears

Background: There are 600,000 new malaria cases daily worldwide. The gold standard for estimating the parasite burden and the corresponding severity of the disease consists in manually counting the number of parasites in blood smears through a microscope, a process that can take more than 20 minutes of an expert microscopist’s time. Objective: This research tests the feasibility of a crowdsourced approach to malaria image analysis. In particular, we investigated whether anonymous volunteers with no prior experience would be able to count malaria parasites in digitized images of thick blood smears by playing a Web-based game. Methods: The experimental system consisted of a Web-based game where online volunteers were tasked with detecting parasites in digitized blood sample images coupled with a decision algorithm that combined the analyses from several players to produce an improved collective detection outcome. Data were collected through the MalariaSpot website. Random images of thick blood films containing Plasmodium falciparum at medium to low parasitemias, acquired by conventional optical microscopy, were presented to players. In the game, players had to find and tag as many parasites as possible in 1 minute. In the event that players found all the parasites present in the image, they were presented with a new image. In order to combine the choices of different players into a single crowd decision, we implemented an image processing pipeline and a quorum algorithm that judged a parasite tagged when a group of players agreed on its position. Results: Over 1 month, anonymous players from 95 countries played more than 12,000 games and generated a database of more than 270,000 clicks on the test images. Results revealed that combining 22 games from nonexpert players achieved a parasite counting accuracy higher than 99%. This performance could be obtained also by combining 13 games from players trained for 1 minute. Exhaustive computations measured the parasite counting accuracy for all players as a function of the number of games considered and the experience of the players. In addition, we propose a mathematical equation that accurately models the collective parasite counting performance. Conclusions: This research validates the online gaming approach for crowdsourced counting of malaria parasites in images of thick blood films. The findings support the conclusion that nonexperts are able to rapidly learn how to identify the typical features of malaria parasites in digitized thick blood samples and that combining the analyses of several users provides similar parasite counting accuracy rates as those of expert microscopists. This experiment illustrates the potential of the crowdsourced gaming approach for performing routine malaria parasite quantification, and more generally for solving biomedical image analysis problems, with future potential for telediagnosis related to global health challenges

Implications for phase separation

Funding Information: The authors would also like to acknowledge Prof. Dr. Jaime Mota, Dra. Irina Franco for the technical assistance with the microscopic experiments, Philip O'Toole for the aid in protein production and Dr. Aldino Viegas and Dr. David Pantoja-Uceda for the support and valuable discussions regarding NMR spectroscopy. This work was supported by Fundação para a Ciência e a Tecnologia (FCT-Portugal) for funding UCIBIO project (UIDP/04378/2020 and UIDB/04378/2020) and Associate Laboratory Institute for Health and Bioeconomy – i4HB Project (LA/P/0140/2020). The authors also thank FCT-Portugal for the PhD grant attributed to SF (PD/BD/148028/2019) under the PTNMRPhD Program. JO is a recipient of a Leonardo Grant from the Spanish BBVA Foundation (BBM_TRA_0203) and a Ramón y Cajal Grant (RYC2018-026042-I funded by MCIN/AEI/10.13039/501100011033 and by “ESF Investing in your future.”) JO and DVL are supported by the Spanish Grants PID-2019-109276RA-I00 and PID-2019-109306RB-I00, respectively, both funded by MCIN/AEI/10.13039/501100011033. The NMR spectrometers are part of the National NMR Facility supported by FCT-Portugal (ROTEIRO/0031/2013–PINFRA/22161/2016, co-financed by FEDER through COMPETE 2020, POCI and PORL and FCT through PIDDAC). The 800 MHz spectrometer present in the “Manuel Rico” NMR laboratory (LMR-CSIC) is a node of the Spanish Large-Scale National Facility (ICTS R-LRB-MR). Publisher Copyright: © 2022 The Authors. Protein Science published by Wiley Periodicals LLC on behalf of The Protein Society.The mediation of liquid–liquid phase separation (LLPS) for fused in sarcoma (FUS) protein is generally attributed to the low-complexity, disordered domains and is enhanced at low temperature. The role of FUS folded domains on the LLPS process remains relatively unknown since most studies are mainly based on fragmented FUS domains. Here, we investigate the effect of metabolites on full-length (FL) FUS LLPS using turbidity assays and differential interference contrast (DIC) microscopy, and explore the behavior of the folded domains by nuclear magnetic resonance (NMR) spectroscopy. FL FUS LLPS is maximal at low concentrations of glucose and glutamate, moderate concentrations of NaCl, Zn2+, and Ca2+ and at the isoelectric pH. The FUS RNA recognition motif (RRM) and zinc-finger (ZnF) domains are found to undergo cold denaturation above 0°C at a temperature that is determined by the conformational stability of the ZnF domain. Cold unfolding exposes buried nonpolar residues that can participate in LLPS-promoting hydrophobic interactions. Therefore, these findings constitute the first evidence that FUS globular domains may have an active role in LLPS under cold stress conditions and in the assembly of stress granules, providing further insight into the environmental regulation of LLPS.publishersversionpublishe

Structural transitions in orb2 prion-like domain relevant for functional aggregation in memory consolidation

Grant BBM_TRA_0203 PD/BD/148028/2019 UIDB/04378/2020The recent structural elucidation of ex vivo Drosophila Orb2 fibrils revealed a novel amyloid formed by interdigitated Gln and His residue side chains belonging to the prion-like domain. However, atomic-level details on the conformational transitions associated with memory consolidation remain unknown. Here, we have characterized the nascent conformation and dynamics of the prion-like domain (PLD) of Orb2A using a nonconventional liquid-state NMR spectroscopy strategy based on 13C detection to afford an essentially complete set of 13Ca, 13Cb, 1Ha, and backbone 13CO and 15N assignments. At pH 4, where His residues are protonated, the PLD is disordered and flexible, except for a partially populated a-helix spanning residues 55–60, and binds RNA oligos, but not divalent cations. At pH 7, in contrast, His residues are predominantly neutral, and the Q/H segments adopt minor populations of helical structure, show decreased mobility and start to self-associate. At pH 7, the His residues do not bind RNA or Ca21, but do bind Zn21, which promotes further association. These findings represent a remarkable case of structural plasticity, based on which an updated model for Orb2A functional amyloidogenesis is suggested.publishersversionpublishe

Atlas Toolkit: Fast registration of 3D morphological datasets in the absence of landmarks

Image registration is a gateway technology for Developmental Systems Biology, enabling computational analysis of related datasets within a shared coordinate system. Many registration tools rely on landmarks to ensure that datasets are correctly aligned; yet suitable landmarks are not present in many datasets. Atlas Toolkit is a Fiji/ImageJ plugin collection offering elastic group-wise registration of 3D morphological datasets, guided by segmentation of the interesting morphology. We demonstrate the method by combinatorial mapping of cell signalling events in the developing eyes of chick embryos, and use the integrated datasets to predictively enumerate Gene Regulatory Network states