SARS-CoV-2 uses CD4 to infect T helper lymphocytes.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the agent of a major global outbreak of respiratory tract disease known as Coronavirus Disease 2019 (COVID-19). SARS-CoV-2 infects mainly lungs and may cause several immune-related complications, such as lymphocytopenia and cytokine storm, which are associated with the severity of the disease and predict mortality. The mechanism by which SARS-CoV-2 infection may result in immune system dysfunction is still not fully understood. Here, we show that SARS-CoV-2 infects human CD4+ T helper cells, but not CD8+ T cells, and is present in blood and bronchoalveolar lavage T helper cells of severe COVID-19 patients. We demonstrated that SARS-CoV-2 spike glycoprotein (S) directly binds to the CD4 molecule, which in turn mediates the entry of SARS- CoV-2 in T helper cells. This leads to impaired CD4 T cell function and may cause cell death. SARS-CoV-2-infected T helper cells express higher levels of IL-10, which is associated with viral persistence and disease severity. Thus, CD4-mediated SARS-CoV-2 infection of T helper cells may contribute to a poor immune response in COVID-19 patients.

Structural discrimination analysis for constraint selection in protein modeling.

MOTIVATION: Protein structure modeling can be improved by the use of distance constraints between amino acid residues, provided such data reflects-at least partially-the native tertiary structure of the target system. In fact, only a small subset of the native contact map is necessary to successfully drive the model conformational search, so one important goal is to obtain the set of constraints with the highest true-positive rate, lowest redundancy and greatest amount of information. In this work, we introduce a constraint evaluation and selection method based on the point-biserial correlation coefficient, which utilizes structural information from an ensemble of models to indirectly measure the power of each constraint in biasing the conformational search toward consensus structures. RESULTS: Residue contact maps obtained by direct coupling analysis are systematically improved by means of discriminant analysis, reaching in some cases accuracies often seen only in modern deep-learning-based approaches. When combined with an iterative modeling workflow, the proposed constraint classification optimizes the selection of the constraint set and maximizes the probability of obtaining successful models. The use of discriminant analysis for the valorization of the information of constraint datasets is a general concept with possible applications to other constraint types and modeling problems. AVAILABILITY AND IMPLEMENTATION: MSA for the targets in this work is available on https://github.com/m3g/2021_Bottino_Biserial. Modeling data supporting the findings of this study was generated at the Center for Computing in Engineering and Sciences, and is available from the corresponding author LM on request. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

SARS-CoV-2 Uses CD4 to Infect T Helper Lymphocytes

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the agent of a major global outbreak of respiratory tract disease known as coronavirus disease-2019 (COVID-19). SARS-CoV-2 infects the lungs and may cause several immune-related complications such as lymphocytopenia and cytokine storm which are associated with the severity of the disease and predict mortality . The mechanism by which SARS-CoV-2 infection may result in immune system dysfunction is not fully understood. Here we show that SARS-CoV-2 infects human CD4+ T helper cells, but not CD8+ T cells, and is present in blood and bronchoalveolar lavage T helper cells of severe COVID-19 patients. We demonstrated that SARS-CoV-2 spike glycoprotein (S) directly binds to the CD4 molecule, which in turn mediates the entry of SARS- CoV-2 in T helper cells in a mechanism that also requires ACE2 and TMPRSS2. Once inside T helper cells, SARS-CoV-2 assembles viral factories, impairs cell function and may cause cell death. SARS-CoV-2 infected T helper cells express higher amounts of IL-10, which is associated with viral persistence and disease severity. Thus, CD4-mediated SARS-CoV-2 infection of T helper cells may explain the poor adaptive immune response of many COVID- 19 patients.

Unveiling the interaction between the molecular motor Myosin Vc and the small GTPase Rab3A.

Vertebrates usually have three class V myosin paralogues (MyoV) to control membrane trafficking in the actin-rich cell cortex, but their functional overlapping or differentiation through cargoes selectivity is yet only partially understood. In this work, we reveal that the globular tail domain of MyoVc binds to the active form of small GTPase Rab3A with nanomolar affinity, a feature shared with MyoVa but not with MyoVb. Using molecular docking analyses guided by chemical cross-linking restraints, we propose a model to explain how Rab3A selectively recognizes MyoVa and MyoVc via a distinct binding site from that used by Rab11A. The MyoVa/c binding interface involves multiple residues from both lobules (I and II) and the short helix at the α2-α3 link region, which is conserved between MyoVa and MyoVc, but not in MyoVb. This motif is also responsible for the selective binding of RILPL2 by MyoVa and potentially MyoVc. Together, these findings support the selective recruitment of MyoVa and MyoVc to exocytic pathways via Rab3A and expand our knowledge about the functional evolution of class V myosins. SIGNIFICANCE: Hormone secretion, neurotransmitter release, and cytoplasm membrane recycling are examples of processes that rely on the interaction of molecular motors and Rab GTPases to regulate the intracellular trafficking and tethering of vesicles. Defects in these proteins may cause neurological impairment, immunodeficiency, and other severe disorders, being fatal in some cases. Despite their crucial roles, little is known about how these molecular motors are selectively recruited by specific members of the large family of Rab GTPases. In this study, we unveil the interaction between the actin-based molecular motor Myosin Vc and the small GTPase Rab3A, a key coordinator of vesicle trafficking and exocytosis in mammalian cells. Moreover, we propose a model for their recognition and demonstrate that Rab3A specifically binds to the globular tail of Myosins Va and Vc, but not of Myosin Vb, advancing our knowledge about the molecular basis for the selective recruitment of class V myosins by Rab GTPases.

Navigating the complexities of coordinated conservation along the river Nile.

The river Nile flows across 11 African countries, supporting millions of human livelihoods, and holding globally important biodiversity and endemism yet remains underprotected. No basin-wide spatial conservation planning has been attempted to date, and the importance of coordinated conservation planning for the Nile's biodiversity remains unknown. We address these gaps by creating a basin-wide conservation plan for the Nile's freshwater fish. We identify priority areas for conservation action and compare cross-boundary collaboration scenarios for achieving biodiversity conservation targets, accounting for river connectivity. We found that collaborative conservation efforts are crucial for reducing conservation costs, saving 34% of costs compared to an uncoordinated, business-as-usual scenario. While most Nile basin countries benefit from coordinating conservation planning, costs and benefits are unequally distributed. We identify "hot spots" consistently selected as conservation priority areas across all collaboration scenarios, and provide a framework for improving return on conservation investment for large and complex river systems globally.

Statistical force-field for structural modeling using chemical cross-linking/mass spectrometry distance constraints.

MOTIVATION: Chemical cross-linking/mass spectrometry (XLMS) is an experimental method to obtain distance constraints between amino acid residues which can be applied to structural modeling of tertiary and quaternary biomolecular structures. These constraints provide, in principle, only upper limits to the distance between amino acid residues along the surface of the biomolecule. In practice, attempts to use of XLMS constraints for tertiary protein structure determination have not been widely successful. This indicates the need of specifically designed strategies for the representation of these constraints within modeling algorithms. RESULTS: A force-field designed to represent XLMS-derived constraints is proposed. The potential energy functions are obtained by computing, in the database of known protein structures, the probability of satisfaction of a topological cross-linking distance as a function of the Euclidean distance between amino acid residues. First, the strategy suggests that XL constraints should be set to shorter distances than usually assumed. Second, the complete statistical force-field improves the models obtained and can be easily incorporated into current modeling methods and software. The force-field was implemented and is distributed to be used within the Rosetta ab initio relax protocol. AVAILABILITY AND IMPLEMENTATION: Force-field parameters and usage instructions are freely available online (http://m3g.iqm.unicamp.br/topolink/xlff). SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

TopoLink: evaluation of structural models using chemical crosslinking distance constraints.

SUMMARY: A software was developed to evaluate structural models using chemical crosslinking experiments. The user provides the types of linkers used and their reactivity, and the observed crosslinks and dead-ends. The software computes the minimum length of a physically inspired linker that connects the reactive atoms of interest, and reports the consistency of each distance with the experimental observation. Statistics on model consistency with the links are provided. Tools to evaluate the correlation of crosslinks in ensembles of models were developed. TopoLink was used to evaluate the potential crosslinks of all structures of the CATH database. The number of crosslinks expected as a function of protein size and linker length can be used as guide for experimental design. AVAILABILITY AND IMPLEMENTATION: TopoLink is available as free software at http://m3g.iqm.unicamp.br/topolink, and distributed as source code with a user-friendly graphical interface for Windows. A web server is also provided. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Enhancing protein fold determination by exploring the complementary information of chemical cross-linking and coevolutionary signals.

Motivation: Elucidation of protein native states from amino acid sequences is a primary computational challenge. Modern computational and experimental methodologies, such as molecular coevolution and chemical cross-linking mass-spectrometry allowed protein structural characterization to previously intangible systems. Despite several independent successful examples, data from these distinct methodologies have not been systematically studied in conjunction. One challenge of structural inference using coevolution is that it is limited to sequence fragments within a conserved and unique domain for which sufficient sequence datasets are available. Therefore, coupling coevolutionary data with complimentary distance constraints from orthogonal sources can provide additional precision to structure prediction methodologies. Results: In this work, we present a methodology to combine residue interaction data obtained from coevolutionary information and cross-linking/mass spectrometry distance constraints in order to identify functional states of proteins. Using a combination of structure-based models (SBMs) with optimized Gaussian-like potentials, secondary structure estimation and simulated annealing molecular dynamics, we provide an automated methodology to integrate constraint data from diverse sources in order to elucidate the native conformation of full protein systems with distinct complexity and structural topologies. We show that cross-linking mass spectrometry constraints improve the structure predictions obtained from SBMs and coevolution signals, and that the constraints obtained by each method have a useful degree of complementarity that promotes enhanced fold estimates. Availability and implementation: Scripts and procedures to implement the methodology presented herein are available at https://github.com/mcubeg/DCAXL. Supplementary information: Supplementary data are available at Bioinformatics online.

AgNPs: The New Allies Against S. Mutans Biofilm - A Pilot Clinical Trial and Microbiological Assay.

The purpose of this study was to evaluate the antimicrobial properties of a new formulation containing silver nanoparticles, named Nano Silver Fluoride (NSF), to inhibit Streptococcus mutans biofilm formation on children's dental enamel. The variations in dental biofilm pH and in the Simplified-Oral-Hygiene-Index (OHI-S) also were evaluated after the treatment with NSF. This was a randomized, double-blind, crossover and prospective pilot clinical trial study in which 12 schoolchildren, aged between 7-8 years, had their dental enamel treated with two solutions: S1 - Nano Silver Fluoride and S2 - negative control (saline solution), in different experimental moments. The dental biofilm adhered to enamel treated with NSF had lower values of S. mutans viability (absorbance) and colony forming units (CFU) than the S0 (baseline) and S2. There was a statistically significant difference between the OHI-S mean values of S0 and S1. There were no differences between the biofilm pH (both before and after the use of the test substances) and among the different groups. These properties suggest that NSF has bactericidal effect against S. mutans biofilm and it may be used for clinical control and prevention of dental biofilm formation.

Antimicrobial and cytotoxicity evaluation of colloidal chitosan - silver nanoparticles - fluoride nanocomposites.

The present study aimed to evaluate the antimicrobial activity and cytotoxicity of colloidal chitosan - silver nanoparticle - fluoride nanocomposites (CChAgNpFNc), with different silver nanoparticle shapes and sizes. The syntheses of CChAgNpFNc were performed with silver nitrate added to a chitosan solution, addition of a sodium borohydride solution and solid sodium fluoride. Solution of ascorbic acid was added to synthesize larger silver nanoparticles. CChAgNpFNc obtained: S1- 100% spherical, 8.7±3.1nm; S2- 97% spherical, 15.0±7.9nm and 2.5% triangular, 22.2±9.5nm; S3- 77.3% spherical, 31.8±10.4nm, 15.9% triangular, 27.1±10.1nm and 6.8% elliptical, 33.2±7.8nm; and S4- 75.2% spherical, 43.2±14.3nm; 23.3% triangular 38.2±14.8nm, and 1.5% elliptical 38.4±11.6nm. The CChAgNpFNc showed antimicrobial activity against Staphylococcus aureus, Escherichia coli, Enterococcus faecalis, Pseudomonas aeruginosa and Candida albicans, by microdilution technique. The influence on the growth of microorganisms was evaluated using a fluorescence assay, and showed an increasing lag phase and a decreasing log phase. Cytotoxicity was investigated using Artemia salina and MTT assays. The S3 and S4 samples exhibited low cytotoxicity. The S1 and S2 samples inhibited murine macrophages and revealed lethal dose concentrations above 1000mg/mL that were classified as moderately toxic. Thus, CChAgNpFNc are potential options for the control of multiple-drug-resistant microorganisms and do not represent substantial risks to human health.

Crystal structure of the human Tip41 orthologue, TIPRL, reveals a novel fold and a binding site for the PP2Ac C-terminus.

TOR signaling pathway regulator-like (TIPRL) is a regulatory protein which inhibits the catalytic subunits of Type 2A phosphatases. Several cellular contexts have been proposed for TIPRL, such as regulation of mTOR signaling, inhibition of apoptosis and biogenesis and recycling of PP2A, however, the underlying molecular mechanism is still poorly understood. We have solved the crystal structure of human TIPRL at 2.15 Å resolution. The structure is a novel fold organized around a central core of antiparallel beta-sheet, showing an N-terminal α/ß region at one of its surfaces and a conserved cleft at the opposite surface. Inside this cleft, we found a peptide derived from TEV-mediated cleavage of the affinity tag. We show by mutagenesis, pulldown and hydrogen/deuterium exchange mass spectrometry that this peptide is a mimic for the conserved C-terminal tail of PP2A, an important region of the phosphatase which regulates holoenzyme assembly, and TIPRL preferentially binds the unmodified version of the PP2A-tail mimetic peptide DYFL compared to its tyrosine-phosphorylated version. A docking model of the TIPRL-PP2Ac complex suggests that TIPRL blocks the phosphatase's active site, providing a structural framework for the function of TIPRL in PP2A inhibition.

In vitro antimicrobial activity of solution blow spun poly(lactic acid)/polyvinylpyrrolidone nanofibers loaded with Copaiba (Copaifera sp.) oil.

In this study poly(lactic acid) (PLA) and polyvinylpyrrolidone (PVP) micro- and nanofiber mats loaded with Copaiba (Copaifera sp.) oil were produced by solution blow spinning (SBS). The Copaiba (Copaifera sp.) oil was characterized by gas chromatography (GC). Neat PLA and four PLA/PVP blends containing 20% (wt.%) oil were spun and characterized by scanning electron microscopy (SEM) and by studying the surface contact angle, in vitro release rate, and antimicrobial activity. All compositions evaluated were able to produce continuous and smooth fibers by SBS. The addition of PVP increased fiber diameter, and decreased the surface contact angle. GC analysis demonstrated that the main component of the Copaiba oil was ß-caryophyllene, a known antimicrobial agent. In vitro release tests of Copaiba oil volatiles demonstrated a higher release rate in fibers containing PVP. Fiber mats made from blends containing higher amounts of PVP had greater antimicrobial action against Staphylococcus aureus. The results confirm the potential of the fiber mats for use in controlled drug release and could lead to promising applications in the biomedical field.

Action of silver nanoparticles towards biological systems: cytotoxicity evaluation using hen's egg test and inhibition of Streptococcus mutans biofilm formation.

This study aimed to evaluate the cytotoxicity and bactericidal properties of four silver nanoparticle (AgNP) colloids and their ability to inhibit Streptococcus mutans biofilm formation on dental enamel. The cytotoxicity of AgNPs was evaluated based on signs of vascular change on the chorioallantoic membrane using the hen's egg test (HET-CAM). Bactericidal properties and inhibition of S. mutans biofilm formation were determined using a parallel-flow cell system and a dichromatic fluorescent stain. The percentage of viable cells was calculated from regression data generated from a viability standard. AgNP colloids proved to be non-irritating, as they were unable to promote vasoconstriction, haemorrhage or coagulation. AgNP colloids inhibited S. mutans biofilm formation on dental enamel, and cell viability measured by fluorescence was 0% for samples S1, S2, S3 and S4 and 36.5% for the positive control (diluted 30% silver diamine fluoride). AgNPs are new products with a low production cost because they have a lower concentration of silver, with low toxicity and an effective bactericidal effect against a cariogenic oral bacterium. Moreover, they do not promote colour change in dental enamel, which is an aesthetic advantage compared with traditional silver products.

Effect of head or neck cooling used with a liquid-conditioned vest during simulated aircraft sorties.

The effectiveness of head or neck cooling in reducing head sweating and increasing subject comfort when used in conjunction with a liquid-conditioned vest during simulated sorties in the European Fighter Aircraft was investigated. Six subjects underwent three 2-h exposures at 40 degrees C wearing Aircrew Chemical Defence clothing and one of three combinations of liquid-conditioned garments. In all three exposures, a liquid-conditioned vest was worn and, in two exposures, either a head-cooling cowl or a neck-cooling collar was worn in addition to the vest. All six subjects reported increased comfort and decreased head sweating with head cooling, a result supported by the data collected.

Detectability of emergency lights for underwater escape.

The time to detect each of three underwater lights by six subjects was measured in clear and turbid (attenuation coefficient = 4.2.m-1) water, at distances of 1.54 m and 3.1 m, from two viewing angles, straight ahead (0 degrees) or 65 degrees to one side, and under three levels of ambient illumination. The lights were viewed either through a window, to simulate the use of a face mask, or with the subjects immersed. All lights were detected rapidly (less than 1 s) when viewed through the window in clear water. In turbid conditions, none of the lights was seen at 3.1 m by any subject. At a 1.54 m viewing distance, reliable detection by immersed subjects was found only in the clear water under the two darker ambient illuminations. In turbid water, detection was unreliable. We conclude that the design of underwater escape lighting should not rely on visibility over distances greater than 1.5 m, and that an illuminated guide-bar might provide valuable assistance in directing escape from aircraft.

The thermal performance of partial coverage wet suits.

A wet-suit worn external to normal clothing and covering the trunk and arms only has been assessed as a method for providing short-term immersion protection for helicopter passengers in offshore oil field operations. Manikin measurements of effective insulation in water give a mean figure of 0.54 togs for the areas covered by the suit and 0.09 togs for uncovered areas. These figures were used to obtain model predictions of survival time for 'thin' and 'average' men which suggest that the suit can give adequate protection for 1 h at 5 degrees C subject to care in fitting. Direct measurements of heat flux have demonstrated the presence of water flushing beneath the suit and the potentially serious loss of insulation that can result.

Immersion cooling: effect of clothing and skinfold thickness.

Accidental immersion often involves the threat of death due to hypothermia. Clothing to control heat loss in water is generally selected to minimize bulk while providing the necessary protection. While water temperature (Tw) and possible immersion time are often considered, another relevant variable is the insulation provided by subcutaneous fat. This paper describes the use of a sophisticated computer model to explore the interactions among skinfold thickness (6-20 mm mean weighted value), clothing insulation (0.06-0.23 clo, immersed), and Tw (0-20 degrees C), in producing critical hypothermia (arterial temperature less than or equal to 34 degrees C). Results indicate that subcutaneous fat strongly affects heat loss even with heavy clothing. Discussion includes examples of the possible use of skinfold data to improve specification of protective clothing for groups and allow special clothing prescription for individuals.

The effect of leakage on the insulation provided by immersion-protection clothing.

The effect of controlled, incremental water leakage on the thermal insulation provided by three immersion-protection assemblies has been measured using a thermal manikin. The results show an average loss of 30% of the initial insulation for a leak of 500 g, 40% for a leak of 1000 g, and nearly 60% for a leak of 3000 g. The assemblies differed only in the thermal insulation layer which consisted of: A. A single thickness of wool (initial immersed insulation 0.63 clo) B. A double thickness of wool (initial immersed insulation 0.79 clo) and C. A layer of polypropylene batting between nylon covers (initial immersed insulation 0.76 clo). Differences between the assemblies in loss of insulation with leakage were small and no evidence was found to support claims that the insulating properties of polypropylene are more resistant to wetting than those of wool. It is considered that the substantial loss of insulation even with small leaks makes it essential that tests of the water-excluding performance of immersion suits are undertaken in realistic conditions rather than in calm water.

Heat stress in front and rear cockpits of F-4 aircraft.

Heat stress can be a serious problem in aircraft flown at low altitudes in warm-to-hot environments. Data are reported here on 36 flights by F-4 aircraft at Eglin AFB, FL. Ground dry-bulb temperatures (Tdb,g) were 19-33 degrees C (mean 28 degrees C) with psychrometric wet-bulb temperatures (Twb) 14-24 degrees C (mean 20 degrees C). Environmental and physiological data were recorded in both front and rear cockpits at 2-min intervals throughout each mission, which simulated low-level ground attack and lasted 94-126 min. Data were analyzed for four phases. A) preflight taxi, B) low-level flight, C) ordnance delivery, and D) postflight taxi. Cockpit dry-bulb temperature (Tdb,c) exceeded Tdb,g during ground operations; the front cockpit cooled in flight, while the rear remained hot. Linear relationships appeared for Tdb,c vs. Tdb,g in the four mission phases, and for globe temperature vs. Tdb,c. Aircrew mean skin temperature was significantly related to Tdb,c, and core (ear canal) temperature rose slightly with heat stress. Sweat rates reflected both Tdb,c and clothing worn. Discussion covers differences in air conditioning in the two cockpits and possible consequences of the observed heat stress and physiological strain.