Modeling the β-secretase cleavage site and humanizing amyloid-beta precursor protein in rat and mouse to study Alzheimer's disease

BACKGROUND: Three amino acid differences between rodent and human APP affect medically important features, including β-secretase cleavage of APP and Aβ peptide aggregation (De Strooper et al., EMBO J 14:4932-38, 1995; Ueno et al., Biochemistry 53:7523-30, 2014; Bush, 2003, Trends Neurosci 26:207-14). Most rodent models for Alzheimer's disease (AD) are, therefore, based on the human APP sequence, expressed from artificial mini-genes randomly inserted in the rodent genome. While these models mimic rather well various biochemical aspects of the disease, such as Aβ-aggregation, they are also prone to overexpression artifacts and to complex phenotypical alterations, due to genes affected in or close to the insertion site(s) of the mini-genes (Sasaguri et al., EMBO J 36:2473-87, 2017; Goodwin et al., Genome Res 29:494-505, 2019). Knock-in strategies which introduce clinical mutants in a humanized endogenous rodent APP sequence (Saito et al., Nat Neurosci 17:661-3, 2014) represent useful improvements, but need to be compared with appropriate humanized wildtype (WT) mice. METHODS: Computational modelling of the human β-CTF bound to BACE1 was used to study the differential processing of rodent and human APP. We humanized the three pivotal residues we identified G676R, F681Y and R684H (labeled according to the human APP770 isoform) in the mouse and rat genomes using a CRISPR-Cas9 approach. These new models, termed mouse and rat Apphu/hu, express APP from the endogenous promotor. We also introduced the early-onset familial Alzheimer's disease (FAD) mutation M139T into the endogenous Rat Psen1 gene. RESULTS: We show that introducing these three amino acid substitutions into the rodent sequence lowers the affinity of the APP substrate for BACE1 cleavage. The effect on β-secretase processing was confirmed as both humanized rodent models produce three times more (human) Aβ compared to the original WT strain. These models represent suitable controls, or starting points, for studying the effect of transgenes or knock-in mutations on APP processing (Saito et al., Nat Neurosci 17:661-3, 2014). We introduced the early-onset familial Alzheimer's disease (FAD) mutation M139T into the endogenous Rat Psen1 gene and provide an initial characterization of Aβ processing in this novel rat AD model. CONCLUSION: The different humanized APP models (rat and mouse) expressing human Aβ and PSEN1 M139T are valuable controls to study APP processing in vivo allowing the use of a human Aβ ELISA which is more sensitive than the equivalent system for rodents. These animals will be made available to the research community

Decreasing population selection rates of resistance mutation K65R over time in HIV-1 patients receiving combination therapy including tenofovir

Objectives The use of tenofovir is highly associated with the emergence of mutation K65R, which confers broad resistance to nucleoside/nucleotide analogue reverse transcriptase inhibitors (NRTIs), especially when tenofovir is combined with other NRTIs also selecting for K65R. Although recent HIV-1 treatment guidelines discouraging these combinations resulted in reduced K65R selection with tenofovir, updated information on the impact of currently recommended regimens on the population selection rate of K65R is presently lacking. Methods In this study, we evaluated changes over time in the selection rate of resistance mutation K65R in a large population of 2736 HIV-1-infected patients failing combination antiretroviral treatment between 2002 and 2010. Results The K65R resistance mutation was detected in 144 patients, a prevalence of 5.3%. A large majority of observed K65R cases were explained by the use of tenofovir, reflecting its wide use in clinical practice. However, changing patterns over time in NRTIs accompanying tenofovir resulted in a persistent decreasing probability of K65R selection by tenofovir-based therapy. The currently recommended NRTI combination tenofovir/emtricitabine was associated with a low probability of K65R emergence. For any given dual NRTI combination including tenofovir, higher selection rates of K65R were consistently observed with a non-nucleoside reverse transcriptase inhibitor than with a protease inhibitor as the third agent. Discussion Our finding of a stable time trend of K65R despite elevated use of tenofovir illustrates increased potency of current HIV-1 therapy including tenofovi

The spatial scale of ozone depletion events derived from an autonomous surface ozone network in coastal Antarctica

To probe the spatial extent of tropospheric ozone depletion events during Antarctic spring, a network of 10 autonomous ozone monitors was established around the Dronning Maud Land sector of Antarctica for a full calendar year. Together with manned stations in the area, the network covered a ~1200 km stretch of coast, as well as a transect ~300 km inland and to ~2000 m above sea level (a.s.l.). Here we present results from the spring period (August to October 2008). While some ozone depletion events were evident at only a single site, implying localised ozone destruction, others were evident across the network. The fact that, on occasions, ozone depletion events were observed at all coastal sites simultaneously, suggests the depleted air mass had a scale of at least 1200 km. As the ozone-poor air was advected from the Weddell Sea sea ice zone, the data imply that large areas over the Weddell Sea sea ice zone are significantly depleted in ozone on occasions during Antarctic spring

HIV-1 gp120 N-linked glycosylation differs between plasma and leukocyte compartments

<p>Abstract</p> <p>Background</p> <p>N-linked glycosylation is a major mechanism for minimizing virus neutralizing antibody response and is present on the Human Immunodeficiency Virus (HIV) envelope glycoprotein. Although it is known that glycosylation changes can dramatically influence virus recognition by the host antibody, the actual contribution of compartmental differences in N-linked glycosylation patterns remains unclear.</p> <p>Methodology and Principal Findings</p> <p>We amplified the <it>env </it>gp120 C2-V5 region and analyzed 305 clones derived from plasma and other compartments from 15 HIV-1 patients. Bioinformatics and Bayesian network analyses were used to examine N-linked glycosylation differences between compartments. We found evidence for cellspecific single amino acid changes particular to monocytes, and significant variation was found in the total number of N-linked glycosylation sites between patients. Further, significant differences in the number of glycosylation sites were observed between plasma and cellular compartments. Bayesian network analyses showed an interdependency between N-linked glycosylation sites found in our study, which may have immense functional relevance.</p> <p>Conclusion</p> <p>Our analyses have identified single cell/compartment-specific amino acid changes and differences in N-linked glycosylation patterns between plasma and diverse blood leukocytes. Bayesian network analyses showed associations inferring alternative glycosylation pathways. We believe that these studies will provide crucial insights into the host immune response and its ability in controlling HIV replication <it>in vivo</it>. These findings could also have relevance in shielding and evasion of HIV-1 from neutralizing antibodies.</p

Ground-based measurements of tropospheric and stratospheric bromine monoxide above Nairobi (1° S, 36° E)

International audienceGround based observations of stratospheric and tropospheric bromine monoxide, BrO, from a multi axial differential optical absorption spectrometer, MAXDOAS, located at the UNEP/UNON site in Nairobi (1° S, 36° E) are presented for the year 2003. Differences in BrO slant column densities at 90° and 80° solar zenith angle retrieved from the zenith-sky measurements are used to study stratospheric BrO. They show only small variations with season, as expected for the small seasonality in stratospheric Bry and NO2 in this region. A pronounced diurnal variation can be observed, the average value for the morning being 1.3×1014 molecules/cm2 and for the evening 1.5×1014 molecules/cm2. The measurements are compared with simulations from a one-dimensional photochemical stacked box model which is coupled with a radiative transfer model to allow direct comparisons between the observations and the model calculations. In general the model reproduces the measurements very well. The differences in the absolute values are 15% for the evening and 20% for the morning which is within the limits of the combined uncertainties. Both seasonality and diurnal variation are well reproduced by the model. A sensitivity study shows that inclusion of the reaction BrONO2 + O(3P) significantly improves the agreement between model calculations and measurements, indicating an important role of this reaction in the stratosphere near to the equator. Tropospheric BrO columns and profile information is derived from the combined results obtained in the different viewing directions for the average over several clear days. The resulting tropospheric BrO columns are in the range of 4?7.5×1012 molecules/cm2 which is significant but lower than in previous studies at mid and high latitudes. The vertical distribution of the tropospheric BrO peaks at about 3 km indicating the absence of local sources at this high altitude site

Assessing transmissibility of HIV-1 drug resistance mutations from treated and from drug-naive individuals (AIDS (2015) 29 (2045-2052))

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On the contribution of Angola to the initial spread of HIV-1

Licenced under CC-BY-NC-NDAngola borders and has long-term links with Democratic Republic of Congo (DRC) as well as high levels of Human Immunodeficiency Virus (HIV) genetic diversity, indicating a potential role in the initial spread of the HIV-1 pandemic. Herein, we analyze 564 C2V3 and 354 pol publicly available sequences from DRC, Republic of Congo (RC) and Angola to better understand the initial spread of the virus in this region. Phylogeographic analyses were performed with the BEAST software. While our results pinpoint the origin of the pandemic to Kinshasa (DRC) around 1906, the introduction of HIV-1 to Angola could have occurred early between the 1910s and 1940s. Furthermore, most of the HIV-1 migrations out of Kinshasa were directed not only to Lubumbashi and Mbuji-Mayi (DRC), but also to Luanda and Brazzaville. Kinshasa census records corroborate these findings, indicating that the early exportation of the virus to Angola might be related to the high number of Angolans in Kinshasa at that time, originated mostly from the North of Angola. In summary, our results place Angola at the epicenter of the early HIV dissemination, together with DRC and RC.info:eu-repo/semantics/publishedVersio

Single-Nucleus RNA-Seq Is Not Suitable for Detection of Microglial Activation Genes in Humans

Single-nucleus RNA sequencing (snRNA-seq) is used as an alternative to single-cell RNA-seq, as it allows transcriptomic profiling of frozen tissue. However, it is unclear whether snRNA-seq is able to detect cellular state in human tissue. Indeed, snRNA-seq analyses of human brain samples have failed to detect a consistent microglial activation signature in Alzheimer's disease. Our comparison of microglia from single cells and single nuclei of four human subjects reveals that, although most genes show similar relative abundances in cells and nuclei, a small population of genes (∼1%) is depleted in nuclei compared to whole cells. This population is enriched for genes previously implicated in microglial activation, including APOE, CST3, SPP1, and CD74, comprising 18% of previously identified microglial-disease-associated genes. Given the low sensitivity of snRNA-seq to detect many activation genes, we conclude that snRNA-seq is not suited for detecting cellular activation in microglia in human disease

Numerical modelling of Auriga's Wheel - a new ring galaxy

We model the formation of Auriga's Wheel - a recently discovered collisional ring galaxy. Auriga's Wheel has a number of interesting features including a bridge of stars linking the neighbouring elliptical to the ring galaxy, and evidence for components of expansion and rotation within the ring. Using N-body/SPH modelling, we study collisions between an elliptical galaxy and a late-type disk galaxy. A near direct collision, with a mildy inclined disk, is found to reasonably reproduce the general system morphology ~50 Myr following the collision. The collision must have a relatively low velocity (initially ~150 km s^{-1}) in order to form the observed bridge, and simultaneously match the galaxies separation. Our best-match model suggests the total disk galaxy is ~5 times more massive than the elliptical. We find that the velocity of expansion of the ring is sensitive to the mass of the elliptical, while insensitive to the encounter velocity. We evolve our simulation beyond the current epoch to study the future destiny of the galaxy pair. In our model, the nucleus moves further away from the plane of the ring in the direction of the stellar bridge. The nucleus eventually merges with the elliptical galaxy ~100 Myr after the present time. The ring continues to expand for ~200 Myr before collapsing back. The low initial relative velocity of the two galaxies will eventually result in a complete merger.Comment: Accepted to MNRAS, March 9th, 2012. 17 pages, 16 figures, no table