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Abstract The performance of Cu-exchanged chabazite (Cu-CHA) for the ammonia-assisted selective catalytic reduction of NO x (NH3-SCR) depends critically on the presence of paired $${[{{{\rm{Cu}}}}{({{{{\rm{NH}}}}}_{3})}_{2}]}^{+}$$ [ Cu ( NH 3 ) 2 ] + complexes. Here, a machine-learning force field augmented with long-range Coulomb interactions is developed to investigate the effect of Al-distribution and Cu-loading on the mobility and pairing of $${[{{{\rm{Cu}}}}{({{{{\rm{NH}}}}}_{3})}_{2}]}^{+}$$ [ Cu ( NH 3 ) 2 ] + complexes. Performing unbiased and constrained molecular dynamics simulations, we obtain unique information inaccessible to first-principle calculations and experiments. The free energy barrier for $${[{{{\rm{Cu}}}}{({{{{\rm{NH}}}}}_{3})}_{2}]}^{+}$$ [ Cu ( NH 3 ) 2 ] + diffusion between CHA-cages depends sensitively on both the local and distant Al-distribution. Importantly, certain Al-distributions and arrangements of neighboring $${[{{{\rm{Cu}}}}{({{{{\rm{NH}}}}}_{3})}_{2}]}^{+}$$ [ Cu ( NH 3 ) 2 ] + and $${{{{{\rm{NH}}}}}_{4}}^{+}$$ NH 4 + cations make paired $${[{{{\rm{Cu}}}}{({{{{\rm{NH}}}}}_{3})}_{2}]}^{+}$$ [ Cu ( NH 3 ) 2 ] + complexes exothermic with respect to separated configurations. Our results suggest that the NH3-SCR activity can be enhanced by increasing the Cu-loading and Al-content. The dynamic interplay between $${[{{{\rm{Cu}}}}{({{{{\rm{NH}}}}}_{3})}_{2}]}^{+}$$ [ Cu ( NH 3 ) 2 ] + and $${{{{{\rm{NH}}}}}_{4}}^{+}$$ NH 4 + diffusion is crucial for the $${[{{{\rm{Cu}}}}{({{{{\rm{NH}}}}}_{3})}_{2}]}^{+}$$ [ Cu ( NH 3 ) 2 ] + mobility and stresses the need to explore large systems including long-range Coulomb interactions when studying diffusion of charged species in zeolites.

Abstract Ultra-fast single-photon detectors with high current density and operating temperature can benefit space and ground applications, including quantum optical communication systems, lightweight cryogenics for space crafts, and medical use. Here we demonstrate magnesium diboride (MgB2) thin-film superconducting microwires capable of single-photon detection at 1.55 μm optical wavelength. We used helium ions to alter the properties of MgB2, resulting in microwire-based detectors exhibiting single-photon sensitivity across a broad temperature range of up to 20 K, and detection efficiency saturation for 1 μm wide microwires at 3.7 K. Linearity of detection rate vs incident power was preserved up to at least 100 Mcps. Despite the large active area of up to 400 × 400 μm2, the reset time was found to be as low as ~ 1 ns. Our research provides possibilities for breaking the operating temperature limit and maximum single-pixel count rate, expanding the detector area, and raises inquiries about the fundamental mechanisms of single-photon detection in high-critical-temperature superconductors.

Abstract Terahertz time-domain spectroscopy (THz-TDS) can be used to map spatial variations in electrical properties such as sheet conductivity, carrier density, and carrier mobility in graphene. Here, we consider wafer-scale graphene grown on germanium by chemical vapor deposition with non-uniformities and small domains due to reconstructions of the substrate during growth. The THz conductivity spectrum matches the predictions of the phenomenological Drude–Smith model for conductors with non-isotropic scattering caused by backscattering from boundaries and line defects. We compare the charge carrier mean free path determined by THz-TDS with the average defect distance assessed by Raman spectroscopy, and the grain boundary dimensions as determined by transmission electron microscopy. The results indicate that even small angle orientation variations below 5° within graphene grains influence the scattering behavior, consistent with significant backscattering contributions from grain boundaries.

Chromolaena odorata (L.) King and Robinson (Asteraceae) is a tropical shrub with interesting chemical potential widely used in agriculture and medical science and which can be affected by several geographic and climatic conditions. Therefore, we investigated the phytochemical composition of this plant across climatic zones in Benin. The plant material collected from different locations was phytochemically screened by staining and precipitation tests. The total phenolic, flavonoid and tannin contents were determined using, the colorimetric method of Folin-Ciocalteu, the method of aluminum chloride and the method of vanillin, respectively, then the obtained data were subjected to analysis of variance. The phytochemical analysis revealed the presence of the main chemical groups such as alkaloids, free anthracene, coumarins, flavonoids, mucilage, tannins, reducing compounds, saponins, quinone derivatives, steroids. There was a significant difference (P < 0.05) in the phytochemical contents across geographical sites. In comparing the levels of phytochemicals among geographical locations, the raw material collected from the north climatic zone contained the highest phenolic and flavonoid contents, 147.59 ± 3.04 mg/g and 17.17 ±0.31 mg/g, respectively, compared to others. Overall, the study highlighted the potential of C. odorata as source of natural products. There was no difference in the phytochemical markers whereas the phytochemical contents vary across climatic zones. These results can be of use in the development of biopesticides from the raw material of C. odorata.

Mutations in the gene encoding dynamin 2 (DNM2), a GTPase that catalyzes membrane constriction and fission, are associated with two autosomal-dominant motor disorders, Charcot-Marie-Tooth disease (CMT) and centronuclear myopathy (CNM), which affect nerve and muscle, respectively. Many of these mutations affect the pleckstrin homology domain of DNM2, yet there is almost no overlap between the sets of mutations that cause CMT or CNM. A subset of CMT-linked mutations inhibit the interaction of DNM2 with phosphatidylinositol (4,5) bisphosphate, which is essential for DNM2 function in endocytosis. In contrast, CNM-linked mutations inhibit intramolecular interactions that normally suppress dynamin self-assembly and GTPase activation. Hence, CNM-linked DNM2 mutants form abnormally stable polymers and express enhanced assembly-dependent GTPase activation. These distinct effects of CMT and CNM mutations are consistent with current findings that DNM2-dependent CMT and CNM are loss-of-function and gain-of-function diseases, respectively. In this study, we present evidence that at least one CMT-causing DNM2 mutant (ΔDEE; lacking residues 555DEE557) forms polymers that, like the CNM mutants, are resistant to disassembly and display enhanced GTPase activation. We further show that the ΔDEE mutant undergoes 2-3-fold higher levels of tyrosine phosphorylation than wild-type DNM2. These results suggest that molecular mechanisms underlying the absence of pathogenic overlap between DNM2-dependent CMT and CNM should be re-examined.

This dataset contains raw area counts and percent recoveries of polycyclic aromatic hydrocarbon (PAH) standards desorbed from selected sorbent tubes and analyzed using thermal desorption-gas chromatography/mass spectrometry (TD-GC/MS). The results of this study were published in the article “Recovery and reactivity of polycyclic aromatic hydrocarbons collected on selected sorbent tubes and analyzed by thermal desorption-gas chromatography/mass spectrometry” in Journal of Chromatography A [1]. The sorbent tubes studied include stainless steel Carbograph 2TD/1TD, glass quartz wool-Carbograph 2TD, inert-coated stainless steel Carbograph 2TD, glass and stainless steel Tenax TA, PAH (chemical weapons), and glass and stainless steel XRO-440 sorbent tubes. Tables listing the experimental conditions, TD methods, and types of sorbent tubes are included in the manuscript. Data for experiments, including the investigation of incomplete desorption of PAHs from Carbograph 2TD/1TD and XRO-440 sorbent tubes, the comparison of PAH recoveries from three different TD methods, the analysis of PAH breakthrough from sorbent tubes, the investigation of the effect of heat on PAH percent recovery from sorbent tubes, and the formation of reaction products during PAH loading and desorption are included in Appendix A. These data can be used to guide sorbent tube selection for PAH analyses in future studies. Keywords: Polycyclic aromatic hydrocarbon (PAH), Thermal desorption, Gas chromatography-mass spectrometry (GC/MS), Sorbent tubes, Carbograph, Tenax, XRO-440

The data contained in this article are PubMed search strings and search string builders used to curate breath research manuscripts published from 1995–2016 and the respective number of articles found that satisfied the search requirements for selected categories. Breath sampling represents a non-invasive technique that has gained usefulness for public health, clinical, diagnostic, and environmental exposure assessment applications over the years. This data article includes search strings that were utilized to retrieve publications through the PubMed database for different breath research-related topics that were related to the analysis of exhaled breath, exhaled breath condensate (EBC), and exhaled breath aerosol (EBA) as well as the analysis of cellular headspace. Manuscripts were curated for topics including EBC, EBA, Direct MS, GC–MS, LC-MS, alcohol, and sensors. A summary of the number of papers published per year for the data retrieved using each of the search strings is also included. These data can be utilized to discern trends in the number of breath research publications in each of the different topics over time. A supplementary Appendix A containing the titles, author lists, journal names, publication dates, PMID numbers, and EntrezUID numbers for each of the journal articles curated using the finalized search strings for the seven breath research-related topics can also be found within this article. The selected manuscripts can be used to explore the impact that breath research has had on expanding the scientific knowledge in each of the investigated topics. Keywords: Exhaled breath, Exhaled breath condensate (EBC), Exhaled breath aerosol (EBA), Mass spectrometry, Sensors, Immunochemistry, Public health, Clinical diagnosis

The exploration of 1D magnetism, frequently portrayed as spin chains, constitutes an actively pursued research field that illuminates fundamental principles in many-body problems and applications in magnonics and spintronics. The inherent reduction in dimensionality often leads to robust spin fluctuations, impacting magnetic ordering and resulting in novel magnetic phenomena. Here, structural, magnetic, and optical properties of highly anisotropic 2D van der Waals antiferromagnets that uniquely host spin chains are explored. First-principle calculations reveal that the weakest interaction is interchain, leading to essentially 1D magnetic behavior in each layer. With the additional degree of freedom arising from its anisotropic structure, the structure is engineered by alloying, varying the 1D spin chain lengths using electron beam irradiation, or twisting for localized patterning, and spin textures are calculated, predicting robust stability of the antiferromagnetic ordering. Comparing with other spin chain magnets, these materials are anticipated to bring fresh perspectives on harvesting low-dimensional magnetism.

Conductance quantization is the hallmark of non-interacting confined systems. The authors show that the quantization in graphene nanoconstrictions with low edge disorder is suppressed in the quantum Hall regime. This is explained by the addition of new conductance channels due to electrostatic screening.

Neuronal defense against oxidative damage is mediated primarily by the glutathione redox system. Traditionally considered a mechanism to protect proteins from irreversible oxidation, mounting evidence supports a role for protein S-glutathionylation in cell signaling in response to changes in intracellular redox status. Here we determined the specific sites on the actin binding protein cofilin that undergo S-glutathionylation. In addition, we show that S-glutathionylation of cofilin reduces its capacity to depolymerize F-actin. We further describe an assay to determine the S-glutathionylation of target proteins in brain tissue from behaving rodents. Using this technique, we show that cofilin in the rat nucleus accumbens undergoes S-glutathionylation during 15-minutes of cued cocaine seeking in the absence of cocaine. Our findings demonstrate that cofilin S-glutathionylation is increased in response to cocaine-associated cues and that increased cofilin S-glutathionylation reduces cofilin-dependent depolymerization of F-actin. Thus, S-glutathionylation of cofilin may serve to regulate actin cycling in response to drug-conditioned cues.

ABSTRACT The assembly of virus particles is a crucial aspect of virus spread. For retroviruses, the Gag polyprotein is the key driver for virus particle assembly. In order to produce progeny virus, once Gag is translated, it must translocate from the location in the cytoplasm where it is synthesized to the plasma membrane and form an oligomeric lattice that results in Gag puncta. The biogenesis of mature Gag puncta can trigger the budding process, resulting in virus particle production. While some aspects of the dynamics of Gag oligomerization and particle biogenesis have been observed with human immunodeficiency virus type 1 (HIV-1), the process of Gag punctum biogenesis remains poorly understood, particularly for other retroviruses. Here, we have conducted the most detailed studies thus far on Gag punctum biogenesis for human T-cell leukemia virus type 1 (HTLV-1). Using mEos2 photoconvertible fluorescent proteins and total internal reflection fluorescence microscopy (TIRF), we have found that HTLV-1 Gag was recruited to Gag puncta primarily from the plasma membrane. This was in stark contrast to HIV-1 Gag, which was recruited from the cytoplasm. These observations imply fundamental differences among retroviruses regarding the orchestration of Gag punctum biogenesis, which has important general implications for enveloped virus particle assembly. IMPORTANCE This report describes the results of experiments examining the pathway by which the human retroviral Gag protein is recruited to sites along the inner leaflet of the plasma membrane where Gag punctum biogenesis occurs. In particular, clever and sensitive experimental methods were devised to image in living cells fluorescently labeled Gag protein derivatives from human T-cell leukemia virus type 1 (HTLV-1) and human immunodeficiency virus type 1 (HIV-1) at the plasma membrane. The photoconvertible fluorescent protein mEos2 was strategically utilized, as the fluorescence emission of Gag at the plasma membrane could be differentiated from that of cytosolic Gag. This experimental strategy allowed for the determination of the Gag recruitment pathway into Gag puncta. For HTLV-1 Gag, puncta recruited Gag primarily from the plasma membrane, while HIV-1 Gag was recruited from the cytoplasm. These observations represent the first report of HTLV-1 particle biogenesis and its contrast to that of HIV-1. The observed differences in the Gag recruitment pathways used by HTLV-1 and HIV-1 Gag provide key information that is useful for informing the discovery of novel targets for antiretroviral therapies directed at eliminating virus infectivity and spread.

We present here a longitudinal study determining the effects of two 3 week-periods of high intensity high volume interval training (HIHVT) (90 intervals of 6 s cycling at 250% maximum power, Pmax/24 s) on a cycle ergometer. HIHVT was evaluated by comparing performance tests before and after the entire training (baseline, BSL, and endpoint, END) and between the two training sets (intermediate, INT). The mRNA expression levels of myosin heavy chain (MHC) isoforms and markers of energy metabolism were analyzed in M. vastus lateralis biopsies by quantitative real-time PCR. In incremental tests peak power (Ppeak) was increased, whereas V˙O2peak was unaltered. Prolonged time-to-exhaustion was found in endurance tests with 65 and 80% Pmax at INT and END. No changes in blood levels of lipid metabolites were detected. Training-induced decreases of hematocrit indicate hypervolemia. A shift from slow MHCI/β to fast MHCIIa mRNA expression occurred after the first and second training set. The mRNA expression of peroxisome proliferator-activated receptor gamma coactivator 1α (PGC-1α), a master regulator of oxidative energy metabolism, decreased after the second training set. In agreement, a significant decrease was also found for citrate synthase mRNA after the second training set, indicating reduced oxidative capacity. However, mRNA expression levels of glycolytic marker enzyme glyceraldehyde-3-phosphate dehydrogenase did not change after the first and second training set. HIHVT induced a nearly complete slow-to-fast fiber type transformation on the mRNA level, which, however, cannot account for the improvements of performance parameters. The latter might be explained by the well-known effects of hypervolemia on exercise performance.

Retrovirus morphogenesis entails assembly of Gag proteins and the viral genome on the host plasma membrane, acquisition of the viral membrane and envelope proteins through budding, and formation of the core through the maturation process. Although in both immature and mature retroviruses, Gag and capsid proteins are organized as paracrystalline structures, the curvatures of these protein arrays are evidently not uniform within one or among all virus particles. The heterogeneity of retroviruses poses significant challenges to studying the protein contacts within the Gag and capsid lattices. This review focuses on current understanding of the molecular organization of retroviruses derived from the sub-nanometer structures of immature virus particles, helical capsid protein assemblies and soluble envelope protein complexes. These studies provide insight into the molecular elements that maintain the stability, flexibility and infectivity of virus particles. Also reviewed are morphological studies of retrovirus budding, maturation, infection and cell-cell transmission, which inform the structural transformation of the viruses and the cells during infection and viral transmission, and lead to better understanding of the interplay between the functioning viral proteins and the host cell.

We have studied cardiac and respiratory functions of aquaporin- 1-deficient mice by the Pressure-Volume-loop technique and by blood gas analysis. In addition, the morphological properties of the animals’ hearts were analysed. In anesthesia under maximal dobutamine stimulation, the mice exhibit a moderately elevated heart rate of < 600 min-1 and an O2 consumption of ~0.6 ml/min/g, which is about twice the basal rate. In this state, which is similar to the resting state of the conscious animal, all cardiac functions including stroke volume and cardiac output exhibited resting values and were identical between deficient and wildtype animals. Likewise, pulmonary and peripheral exchange of O2 and CO2 were normal. In contrast, several morphological parameters of the heart tissue of deficient mice were altered: 1) left ventricular wall thickness was reduced by 12%, 2) left ventricular mass, normalized to tibia length, was reduced by 10-20%, 3) cardiac muscle fiber cross sectional area was decreased by 17%, and 4) capillary density was diminished by 10%. As the P-V-loop technique yielded normal end-diastolic and end-systolic left ventricular volumes, the deficient hearts are characterized by thin ventricular walls in combination with normal intraventricular volumes. The aquaporin-1-deficient heart thus seems to be at a disadvantage compared to the wildtype heart by a reduced left-ventricular wall thickness and an increased diffusion distance between blood capillaries and muscle mitochondria. While under the present quasi-resting conditions these morphological alterations have no consequences for cardiac function, we expect that the deficient hearts will show a reduced maximal cardiac output.

Human T-cell leukemia virus type 1 (HTLV-1) has a reputation for being extremely difficult to study in cell culture. The challenges in propagating HTLV-1 has prevented a rigorous analysis of how these viruses replicate in cells, including the detailed steps involved in virus assembly. The details for how retrovirus particle assembly occurs are poorly understood, even for other more tractable retroviral systems. Recent studies on HTLV-1 using state-of-the-art cryo-electron microscopy and fluorescence-based biophysical approaches explored questions related to HTLV-1 particle size, Gag stoichiometry in virions, and Gag-Gag interactions in living cells. These results provided new and exciting insights into fundamental aspects of HTLV-1 particle assembly—which are distinct from those of other retroviruses, including HIV-1. The application of these and other novel biophysical approaches promise to provide exciting new insights into HTLV-1 replication.

Epidemiological observations of urban inhalation exposures to diesel exhaust (DE) and ozone (O3) have shown pre-clinical cardiopulmonary responses in humans. Identifying the key biological mechanisms that initiate these health bioindicators is difficult due to variability in environmental exposure in time and from person to person. Previously, environmentally controlled human exposure chambers have been used to study DE and O3 dose-response patterns separately, but investigation of co-exposures has not been performed under controlled conditions. Because a mixture is a more realistic exposure scenario for the general public, in this study we investigate the relationships of urban levels of urban-level DE exposure (300 μg/m3), O3 (0.3 ppm), DE + O3 co-exposure, and innate immune system responses. Fifteen healthy human volunteers were studied for changes in ten inflammatory cytokines (interleukins 1β, 2, 4, 5, 8, 10, 12p70 and 13, IFN-γ, and TNF-α) and counts of three white blood cell types (lymphocytes, monocytes, and neutrophils) following controlled exposures to DE, O3, and DE+O3. The results show subtle cytokines responses to the diesel-only and ozone-only exposures, and that a more complex (possibly synergistic) relationship exists in the combination of these two exposures with suppression of IL-5, IL-12p70, IFN-γ, and TNF-α that persists up to 22-hours for IFN-γ and TNF-α. The white blood cell differential counts showed significant monocyte and lymphocyte decreases and neutrophil increases following the DE + O3 exposure; lymphocytes and neutrophils changes also persist for at least 22-hours. Because human studies must be conducted under strict safety protocols at environmental levels, these effects are subtle and are generally only seen with detailed statistical analysis. This study indicates that the observed associations between environmental exposures and cardiopulmonary effects are possibly mediated by inflammatory response mechanisms.

The brightness measured by fluorescence fluctuation spectroscopy specifies the average stoichiometry of a labeled protein in a sample. Here we extended brightness analysis, which has been mainly applied in eukaryotic cells, to prokaryotic cells with E. coli serving as a model system. The small size of the E. coli cell introduces unique challenges for applying brightness analysis that are addressed in this work. Photobleaching leads to a depletion of fluorophores and a reduction of the brightness of protein complexes. In addition, the E. coli cell and the point spread function of the instrument only partially overlap, which influences intensity fluctuations. To address these challenges we developed MSQ analysis, which is based on the mean Q-value of segmented photon count data, and combined it with the analysis of axial scans through the E. coli cell. The MSQ method recovers brightness, concentration, and diffusion time of soluble proteins in E. coli. We applied MSQ to measure the brightness of EGFP in E. coli and compared it to solution measurements. We further used MSQ analysis to determine the oligomeric state of nuclear transport factor 2 labeled with EGFP expressed in E. coli cells. The results obtained demonstrate the feasibility of quantifying the stoichiometry of proteins by brightness analysis in a prokaryotic cell.

Kola nut is chewed in many West African cultures and is used ceremonially. The aim of this study is to investigate some biological effects of Cola nitida’s bark after phytochemical screening. The bark was collected, dried, and then powdered for the phytochemical screening and extractions. Ethanol and ethyl acetate extracts of C. nitida were used in this study. The antibacterial activity was tested on ten reference strains and 28 meat isolated Staphylococcus strains by disc diffusion method. The antifungal activity of three fungal strains was determined on the Potato-Dextrose Agar medium mixed with the appropriate extract. The antioxidant activity was determined by DPPH and ABTS methods. Our data revealed the presence of various potent phytochemicals. For the reference and meat isolated strains, the inhibitory diameter zone was from 17.5±0.7 mm (C. albicans) to 9.5±0.7 mm (P. vulgaris). The MIC ranged from 0.312 mg/mL to 5.000 mg/mL and the MBC from 0.625 mg/mL to >20 mg/mL. The highest antifungal activity was observed with F. verticillioides and the lowest one with P. citrinum. The two extracts have an excellent reducing free radical activity. The killing effect of A. salina larvae was perceptible at 1.04 mg/mL. The purified extracts of Cola nitida’s bark can be used to hold meat products and also like phytomedicine.

Human T-cell leukemia virus type 1 (HTLV-1) is an important cancer-causing human retrovirus that has infected approximately 15 million individuals worldwide. Many aspects of HTLV-1 replication, including virus particle structure and assembly, are poorly understood. Group-specific antigen (Gag) proteins labeled at the carboxy terminus with a fluorophore protein have been used extensively as a surrogate for fluorescence studies of retroviral assembly. How these tags affect Gag stoichiometry and particle morphology has not been reported in detail. In this study, we used an HTLV-1 Gag expression construct with the yellow fluorescence protein (YFP) fused to the carboxy-terminus as a surrogate for the HTLV-1 Gag-Pol to assess the effects of co-packaging of Gag and a Gag-YFP on virus-like particle (VLP) morphology and analyzed particles by cryogenic transmission electron microscopy (cryo-TEM). Scanning transmission electron microscopy (STEM) and fluorescence fluctuation spectroscopy (FFS) were also used to determine the Gag stoichiometry. We found that ratios of 3:1 (Gag:Gag-YFP) or greater resulted in a particle morphology indistinguishable from that of VLPs produced with the untagged HTLV-1 Gag, i.e., a mean diameter of ~113 nm and a mass of 220 MDa as determined by cryo-TEM and STEM, respectively. Furthermore, FFS analysis indicated that HTLV-1 Gag-YFP was incorporated into VLPs in a predictable manner at the 3:1 Gag:Gag-YFP ratio. Both STEM and FFS analyses found that the Gag copy number in VLPs produced with a 3:1 ratio of Gag:Gag-YFP was is in the range of 1500–2000 molecules per VLP. The observations made in this study indicate that biologically relevant Gag–Gag interactions occur between Gag and Gag-YFP at ratios of 3:1 or higher and create a Gag lattice structure in VLPs that is morphologically indistinguishable from that of VLPs produced with just untagged Gag. This information is useful for the quantitative analysis of Gag–Gag interactions that occur during virus particle assembly and in released immature particles.

PML nuclear bodies (PML-NBs) are enigmatic structures of the cell nucleus that act as key mediators of intrinsic immunity against viral pathogens. PML itself is a member of the E3-ligase TRIM family of proteins that regulates a variety of innate immune signaling pathways. Consequently, viruses have evolved effector proteins to modify PML-NBs; however, little is known concerning structure-function relationships of viral antagonists. The herpesvirus human cytomegalovirus (HCMV) expresses the abundant immediate-early protein IE1 that colocalizes with PML-NBs and induces their dispersal, which correlates with the antagonization of NB-mediated intrinsic immunity. Here, we delineate the molecular basis for this antagonization by presenting the first crystal structure for the evolutionary conserved primate cytomegalovirus IE1 proteins. We show that IE1 consists of a globular core (IE1CORE) flanked by intrinsically disordered regions. The 2.3 Å crystal structure of IE1CORE displays an all α-helical, femur-shaped fold, which lacks overall fold similarity with known protein structures, but shares secondary structure features recently observed in the coiled-coil domain of TRIM proteins. Yeast two-hybrid and coimmunoprecipitation experiments demonstrate that IE1CORE binds efficiently to the TRIM family member PML, and is able to induce PML deSUMOylation. Intriguingly, this results in the release of NB-associated proteins into the nucleoplasm, but not of PML itself. Importantly, we show that PML deSUMOylation by IE1CORE is sufficient to antagonize PML-NB-instituted intrinsic immunity. Moreover, co-immunoprecipitation experiments demonstrate that IE1CORE binds via the coiled-coil domain to PML and also interacts with TRIM5α We propose that IE1CORE sequesters PML and possibly other TRIM family members via structural mimicry using an extended binding surface formed by the coiled-coil region. This mode of interaction might render the antagonizing activity less susceptible to mutational escape.

Human cytomegalovirus (HCMV) infections are life-threating to people with a compromised or immature immune system. Upon adhesion, fusion of the virus envelope with the host cell is initiated. In this step, the viral glycoprotein gB is considered to represent the major fusogen. Here, we present for the first time structural data on the binding of an anti-herpes virus antibody and describe the atomic interactions between the antigenic domain Dom-II of HCMV gB and the Fab fragment of the human antibody SM5-1. The crystal structure shows that SM5-1 binds Dom-II almost exclusively via only two CDRs, namely light chain CDR L1 and a 22-residue-long heavy chain CDR H3. Two contiguous segments of Dom-II are targeted by SM5-1, and the combining site includes a hydrophobic pocket on the Dom-II surface that is only partially filled by CDR H3 residues. SM5-1 belongs to a series of sequence-homologous anti-HCMV gB monoclonal antibodies that were isolated from the same donor at a single time point and that represent different maturation states. Analysis of amino acid substitutions in these antibodies in combination with molecular dynamics simulations show that key contributors to the picomolar affinity of SM5-1 do not directly interact with the antigen but significantly reduce the flexibility of CDR H3 in the bound and unbound state of SM5-1 through intramolecular side chain interactions. Thus, these residues most likely alleviate unfavorable binding entropies associated with extra-long CDR H3s, and this might represent a common strategy during antibody maturation. Models of entire HCMV gB in different conformational states hint that SM5-1 neutralizes HCMV either by blocking the pre- to postfusion transition of gB or by precluding the interaction with additional effectors such as the gH/gL complex.

The brightness of fluorescently labeled proteins provides an excellent marker for identifying protein interactions in living cells. Quantitative interpretation of brightness, however, hinges on a detailed understanding of the processes that affect the signal fluctuation of the fluorescent label. Here, we focus on the cumulative influence of photobleaching on brightness measurements in cells. Photobleaching within the finite volume of the cell leads to a depletion of the population of fluorescently labeled proteins with time. The process of photodepletion reduces the fluorescence signal which biases the analysis of brightness data. Our data show that even small reductions in the signal can introduce significant bias into the analysis of the data. We develop a model that quantifies the bias and introduce an analysis method that accurately determines brightness in the presence of photodepletion as verified by experiments with mammalian and yeast cells. In addition, photodepletion experiments with the fluorescent protein EGFP reveal the presence of a photoconversion process, which leads to a marked decrease in the brightness of the EGFP protein. We also identify conditions where the effect of EGFP's photoconversion on brightness experiments can be safely ignored.

Diesel exhaust is a complex mixture of gaseous and particulate compounds resulting from an incomplete combustion of diesel fuel. Controlled human exposures to diesel exhaust and diesel exhaust particles (DEP) have contributed to understanding health effects. Such acute exposure studies of healthy subjects to diesel exhaust and DEP demonstrate a pro-inflammatory effect in the lung and systemically but only at higher concentrations (with a threshold dose approximating 300 µg/m3). Unexpectedly, there appears to be a lack of an inflammatory response to diesel exhaust and DEP in asthmatic individuals. Controlled human exposure studies of cardiovascular effects show that, comparable to other particle-associated exposures, diesel exhaust has a capacity to precipitate coronary artery disease. In addition, there is a relationship between diesel exhaust and DEP exposure and vascular endpoints; these effects in diesel exhaust may be diminished with removal of DEP. Many extra-pulmonary health effects of diesel exhaust exposure, including systemic inflammation, pro-thrombotic changes, and cardiovascular disease, are considered consequent to pro-inflammatory events and inflammation in the lung. Future research will focus on the relative importance of diesel exhaust components, potential interactions between components and other pollutants, effects in sensitive individuals, and effects of longer or repeated exposures.

Human T-cell leukemia virus type 1 (HTLV-1) was the first human retrovirus discovered. Studies on HTLV-1 have been instrumental for our understanding of the molecular pathology of virus-induced cancers. HTLV-1 is the etiological agent of an adult T-cell leukemia (ATL) and can lead to a variety of neurological pathologies, including HTLV-1-associated-myelopathy/tropical spastic paraparesis (HAM/TSP). The ability to treat the aggressive ATL subtypes remains inadequate. HTLV-1 replicates by (1) an infectious cycle involving virus budding and infection of new permissive target cells and (2) mitotic division of cells harboring an integrated provirus. Virus replication initiates host antiviral immunity and the checkpoint control of cell proliferation, but HTLV-1 has evolved elegant strategies to counteract these host defense mechanisms to allow for virus persistence. The study of the molecular biology of HTLV-1 replication has provided crucial information for understanding HTLV-1 replication as well as aspects of viral replication that are shared between HTLV-1 and human immunodeficiency virus type 1 (HIV-1). Here in this review, we discuss the various stages of the virus replication cycle—both foundational knowledge as well as current updates of ongoing research that is important for understanding HTLV-1 molecular pathogenesis as well as in developing novel therapeutic strategies.

The nuclear envelope (NE) consists of two concentric nuclear membranes separated by the lumen, an ∼40-nm-wide fluid layer. NE proteins are implicated in important cellular processes ranging from gene expression to nuclear positioning. Although recent progress has been achieved in quantifying the assembly states of NE proteins in their native environment with fluorescence fluctuation spectroscopy, these studies raised questions regarding the association of NE proteins with nuclear membranes during the assembly process. Monitoring the interaction of proteins with membranes is important because the binding event is often associated with conformational changes that are critical to cellular signaling pathways. Unfortunately, the close physical proximity of both membranes poses a severe experimental challenge in distinguishing luminal and membrane-associated NE proteins. This study seeks to address this problem by introducing new, to our knowledge, fluorescence-based assays that overcome the restrictions imposed by the NE environment. We found that luminal proteins violate the Stokes-Einstein relation, which eliminates a straightforward use of protein mobility as a marker of membrane association within the NE. However, a surprising anomaly in the temperature-dependent mobility of luminal proteins was observed, which was developed into an assay for distinguishing between soluble and membrane-bound NE proteins. We further introduced a second independent tool for distinguishing both protein populations by harnessing the previously reported undulations of the nuclear membranes. These membrane undulations introduce local volume changes that produce an additional fluorescence fluctuation signal for luminal, but not for membrane-bound, proteins. After testing both methods using simple model systems, we apply the two assays to investigate a previously proposed model of membrane association for the luminal domain of SUN2, a constituent protein of the linker of nucleoskeleton and cytoskeleton complex. Finally, we investigate the effect of C- and N-terminal tagging of the luminal ATPase torsinA on its ability to associate with nuclear membranes.

The nucleus is delineated by the nuclear envelope (NE), which is a double membrane barrier composed of the inner and outer nuclear membranes as well as a ∼40-nm wide lumen. In addition to its barrier function, the NE acts as a critical signaling node for a variety of cellular processes, which are mediated by protein complexes within this subcellular compartment. Although fluorescence fluctuation spectroscopy is a powerful tool for characterizing protein complexes in living cells, it was recently demonstrated that conventional fluorescence fluctuation spectroscopy methods are not suitable for applications in the NE because of the presence of slow nuclear membrane undulations. We previously addressed this challenge by developing time-shifted mean-segmented Q (tsMSQ) analysis and applied it to successfully characterize protein homo-oligomerization in the NE. However, many NE complexes, such as the linker of the nucleoskeleton and cytoskeleton complex, are formed by heterotypic interactions, which single-color tsMSQ is unable to characterize. Here, we describe the development of dual-color (DC) tsMSQ to analyze NE heteroprotein complexes built from proteins that carry two spectrally distinct fluorescent labels. Experiments performed on model systems demonstrate that DC tsMSQ properly identifies heteroprotein complexes and their stoichiometry in the NE by accounting for spectral cross talk and local volume fluctuations. Finally, we applied DC tsMSQ to study the assembly of the linker of the nucleoskeleton and cytoskeleton complex, a heteroprotein complex composed of Klarsicht/ANC-1/SYNE homology and Sad1/UNC-84 (SUN) proteins, in the NE of living cells. Using DC tsMSQ, we demonstrate the ability of the SUN protein SUN2 and the Klarsicht/ANC-1/SYNE homology protein nesprin-2 to form a heterocomplex in vivo. Our results are consistent with previously published in vitro studies and demonstrate the utility of the DC tsMSQ technique for characterizing NE heteroprotein complexes.

Self-associating split fluorescent proteins (FPs) are split FPs whose two fragments spontaneously associate to form a functional FP. They have been widely used for labeling proteins, scaffolding protein assembly and detecting cell-cell contacts. Recently developments have expanded the palette of self-associating split FPs beyond the original split GFP1-10/11. However, these new ones have suffered from suboptimal fluorescence signal after complementation. Here, by investigating the complementation process, we have demonstrated two approaches to improve split FPs: assistance through SpyTag/SpyCatcher interaction and directed evolution. The latter has yielded two split sfCherry3 variants with substantially enhanced overall brightness, facilitating the tagging of endogenous proteins by gene editing. Based on sfCherry3, we have further developed a new red-colored trans-synaptic marker called Neuroligin-1 sfCherry3 Linker Across Synaptic Partners (NLG-1 CLASP) for multiplexed visualization of neuronal synapses in living C. elegans, demonstrating its broad applications.

The performance of Cu-exchanged chabazite (Cu-CHA) for the ammonia-assisted selective catalytic reduction of NOx (NH3-SCR) depends critically on the presence of paired [ Cu ( NH 3 ) 2 ] + complexes. Here, a machine-learning force field augmented with long-range Coulomb interactions is developed to investigate the effect of Al-distribution and Cu-loading on the mobility and pairing of [ Cu ( NH 3 ) 2 ] + complexes. Performing unbiased and constrained molecular dynamics simulations, we obtain unique information inaccessible to first-principle calculations and experiments. The free energy barrier for [ Cu ( NH 3 ) 2 ] + diffusion between CHA-cages depends sensitively on both the local and distant Al-distribution. Importantly, certain Al-distributions and arrangements of neighboring [ Cu ( NH 3 ) 2 ] + and NH 4 + cations make paired [ Cu ( NH 3 ) 2 ] + complexes exothermic with respect to separated configurations. Our results suggest that the NH3-SCR activity can be enhanced by increasing the Cu-loading and Al-content. The dynamic interplay between [ Cu ( NH 3 ) 2 ] + and NH 4 + diffusion is crucial for the [ Cu ( NH 3 ) 2 ] + mobility and stresses the need to explore large systems including long-range Coulomb interactions when studying diffusion of charged species in zeolites. Ammonia assisted NOx reduction over Cu-exchanged chabazite requires paired [ Cu ( NH 3 ) 2 ] + complexes. Here, a machine-learning force field is developed to investigate the influence of aluminium distribution on the diffusion mechanisms and pairing of [ Cu ( NH 3 ) 2 ] + complexes. [ABSTRACT FROM AUTHOR]

GABARAP is a member of the ATG8 family of ubiquitin-like autophagy related proteins. It was initially discovered as a facilitator of GABA-A receptor translocation to the plasma membrane and has since been shown to promote the intracellular transport of a variety of other proteins under non-autophagic conditions. We and others have shown that GABARAP interacts with the Type II phosphatidylinositol 4-kinase, PI4K2A, and that this interaction is important for autophagosome-lysosome fusion. Here, we identify a 7-amino acid segment within the PI4K2A catalytic domain that contains the GABARAP interaction motif (GIM). This segment resides in an exposed loop that is not conserved in the other mammalian Type II PI 4-kinase, PI4K2B, explaining the specificity of GABARAP binding to the PI4K2A isoform. Mutation of the PI4K2A GIM inhibits GABARAP binding and PI4K2A-mediated recruitment of cytosolic GABARAP to subcellular organelles. We further show that GABARAP binds to mono-phosphorylated phosphoinositides, PI3P, PI4P, and PI5P, raising the possibility that these lipids contribute to the binding energies that drive GABARAP-protein interactions on membranes. [ABSTRACT FROM AUTHOR]