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Autosomal recessive (AR) complete interferon-gamma receptor 1 (IFN-gamma R1) deficiency, also known as one genetic etiology of Mendelian susceptibility to mycobacterial disease (MSMD), is a life-threatening congenital disease leading to premature death. Affected patients present a pathognomonic predisposition to recurrent and severe infections with environmental mycobacteria or the Mycobacterium bovis bacillus Calmette-Guerin (BCG) vaccine. Current therapeutic options are limited to antibiotic treatment and hematopoietic stem cell transplantation, however with poor outcome. Given the clinical success of gene therapy, we introduce the first lentiviral-based gene therapy approach to restore expression and function of the human IFN-gamma R-downstream signaling cascade. In our study, we developed lentiviral vectors constitutively expressing the human IFN-gamma R1 and demonstrate stable transgene expression without interference with cell viability and proliferation in transduced human hematopoietic cells. Using an IFN-gamma R1-deficient HeLa cell model, we show stable receptor reconstitution and restored IFN-gamma R1 signaling without adverse effect on cell functionality. Transduction of both SV40-immortalized and primary fibroblasts derived from IFN-gamma R1-deficient MSMD patients was able to recover IFN-gamma R1 expression and restore type II IFN signaling upon stimulation with IFN-gamma. In summary, we high-light lentiviral vectors to correct the IFN-gamma mediated immunity and present the first gene therapy approach for patients suffering from AR complete IFN-gamma R1 deficiency.

Opioid use disorder (OUD) is a chronic, relapsing disease. Genetic variability, dysregulated stress system response, and history of opioid experimentation or escalating exposure all contribute to the likelihood of developing OUD, which produces complex brain changes that make it difficult to stop opioid use. Understanding the neurobiology of OUD helps nurses anticipate the behaviors of patients with OUD and approach them with empathy. Here, the authors discuss the pathophysiology of OUD, available screening tools, medical treatments, and behavioral interventions that have demonstrated efficacy in reducing substance use.

Background Due to the affordability of whole-genome sequencing, the genetic association design can now address rare diseases. However, some common statistical association methods only consider homozygosity mapping and need several criteria, such as sliding windows of a given size and statistical significance threshold setting, such asP-value < 0.05 to achieve good power in rare disease association detection. Methods Our region-specific method, called expanded maximal segmental score (eMSS), convertsp-values into continuous scores based on the maximal segmental score (MSS) (Lin et al., 2014) for detecting disease-associated segments. Our eMSS considers the whole genome sequence data, not only regions of homozygosity in candidate genes. Unlike sliding window methods of a given size, eMSS does not need predetermined parameters, such as window size or minimum or maximum number of SNPs in a segment. The performance of eMSS was evaluated by simulations and real data analysis for autosomal recessive diseases multiple intestinal atresia (MIA) and osteogenesis imperfecta (OI), where the number of cases is extremely small. For the real data, the results by eMSS were compared with a state-of-the-art method, HDR-del (Imai et al., 2016). Results Our simulation results show that eMSS had higher power as the number of non-causal haplotype blocks decreased. The type I error for eMSS under different scenarios was well controlled,p< 0.05. For our observed data, the bone morphogenetic protein 1 (BMP1) gene on chromosome 8, the Violaxanthin de-epoxidase-related chloroplast (VDR) gene on chromosome 12 associated with OI, and the tetratricopeptide repeat domain 7A (TTC7A) gene on chromosome 2 associated with MIA have previously been identified as harboring the relevant pathogenic mutations. Conclusions When compared to HDR-del, our eMSS is powerful in analyzing even small numbers of recessive cases, and the results show that the method can further reduce numbers of candidate variants to a very small set of susceptibility pathogenic variants underlying OI and MIA. When we conduct whole-genome sequence analysis, eMSS used 3/5 the computation time of HDR-del. Without additional parameters needing to be set in the segment detection, the computational burden for eMSS is lower compared with that in other region-specific approaches.

Tracking the origin of propagating wave signals in an environment with complex reflective surfaces is, in its full generality, a nearly intractable problem which has engendered multiple domain-specific literatures. We posit that, if the environment and sensor geometries are fixed, machine learning algorithms can "learn" the acoustical geometry of the environment and accurately track signal origin. In this paper, we propose the first machine-learning-based approach to identifying the source locations of semi-stationary, tonal, dolphin-whistle-like sounds in a highly reverberant space, specifically a half-cylindrical dolphin pool. Our algorithm works by supplying a learning network with an overabundance of location "clues", which are then selected under supervised training for their ability to discriminate source location in this particular environment. More specifically, we deliver estimated time-difference-of-arrivals (TDOA's) and normalized cross-correlation values computed from pairs of hydrophone signals to a random forest model for high-feature-volume classification and feature selection, and subsequently deliver the selected features into linear discriminant analysis, linear and quadratic Support Vector Machine (SVM), and Gaussian process models. Based on data from 14 sound source locations and 16 hydrophones, our classification models yielded perfect accuracy at predicting novel sound source locations. Our regression models yielded better accuracy than the established Steered-Response Power (SRP) method when all training data were used, and comparable accuracy along the pool surface when deprived of training data at testing sites; our methods additionally boast improved computation time and the potential for superior localization accuracy in all dimensions with more training data. Because of the generality of our method we argue it may be useful in a much wider variety of contexts.

Atopic dermatitis (AD) is a heterogeneous disease with unique clinical manifestations across age groups and race/ethnicities. Characteristic molecular mechanisms, known as endotypes, including IgE level, status of epidermal barrier genes, and differential cytokine axes activation in the background of T(H)2 upregulation, are also implicated. In adults, the T(H)22, T(H)17, and T(H)1 pathways are involved, and a weakened epidermal barrier is characteristic. In contrast, pediatric patients exhibit less T(H)1 activation, and defects in epidermal lipid metabolism contribute to their barrier defect. European American patients are characterized by higher differential T(H)2/T(H)22 activation, lower expression of the T(H)1/T(H)17 axes, and suppression of filaggrin (FLG) and loricrin gene expressions. Asian patients have accentuated polarity of the T(H)22/T(H)17 pathways, and also exhibit epidermal barrier defects despite relative maintenance of FLG and loricrin expression. African American patients do not exhibit FLG mutations and have distinct attenuation of T(H)17/T(H)1 axes activation. Dissecting the molecular basis of AD endotypes has provided an important framework upon which targeted therapeutics are being developed. An increased understanding of these subtypes and the alteration of biomarkers that correlate with disease can ultimately push AD treatment in an era of personalized medicine. (c) 2020 American Academy of Allergy, Asthma & Immunology

Urinary tract infections (UTIs) typically evoke prompt and vigorous innate bladder immune responses, including extensive exfoliation of the epithelium. To explain the basis for the extraordinarily high recurrence rates of UTIs, we examined adaptive immune responses in mouse bladders. We found that, following each bladder infection, a highly T helper type 2 (T(H)2)-skewed immune response directed at bladder re-epithelialization is observed, with limited capacity to clear infection. This response is initiated by a distinct subset of CD301b(+)OX40L(+) dendritic cells, which migrate into the bladder epithelium after infection before trafficking to lymph nodes to preferentially activate T(H)2 cells. The bladder epithelial repair response is cumulative and aberrant as, after multiple infections, the epithelium was markedly thickened and bladder capacity was reduced relative to controls. Thus, recurrence of UTIs and associated bladder dysfunction are the outcome of the preferential focus of the adaptive immune response on epithelial repair at the expense of bacterial clearance. Abraham and colleagues show that a highly polarized T(H)2 bladder response to urinary tract infections promotes epithelial repair at the expense of preventing new infections and associated bladder dysfunction.

Exocyst is an evolutionarily conserved hetero-octameric tethering complex that plays a variety of roles in membrane trafficking, including exocytosis, endocytosis, autophagy, cell polarization, cytokinesis, pathogen invasion, and metastasis. Exocyst serves as a platform for interactions between the Rab, Rho, and Ral small GTPases, SNARE proteins, and Sec1/Munc18 regulators that coordinate spatial and temporal fidelity of membrane fusion. However, its mechanism is poorly described at the molecular level. Here, we determine the molecular architecture of the yeast exocyst complex by an integrative approach, based on a 3D density map from negative-stain electron microscopy (EM) at similar to 16 angstrom resolution, 434 disuccinimidyl suberate and 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride cross-links from chemical-crosslinking mass spectrometry, and partial atomic models of the eight subunits. The integrative structure is validated by a previously determined cryo-EM structure, cross-links, and distances from in vivo fluorescence microscopy. Our subunit configuration is consistent with the cryo-EM structure, except for Sec5. While not observed in the cryo-EM map, the integrative model localizes the N-terminal half of Sec3 near the Sec6 subunit. Limited proteolysis experiments suggest that the conformation of Exo70 is dynamic, which may have functional implications for SNARE and membrane interactions. This study illustrates how integrative modeling based on varied low-resolution structural data can inform biologically relevant hypotheses, even in the absence of high-resolution data.

In this issue of Cell Chemical Biology, White et al. (2020) describe CRISPR/Cas9-mediated tagging of GPCRs and beta-arrestin to provide a method to study receptor signaling in cells under conditions of endogenous genetic control. The strategy, when coupled with luminescence reporter and complementation technologies, provides new avenues to study GPCRs.

Interferon-gamma receptor 1 (IFN gamma R1) deficiency is one of the inborn errors of IFN-gamma immunity underlying Mendelian Susceptibility to Mycobacterial Disease (MSMD). This molecular circuit plays a crucial role in regulating the interaction between dendritic cells (DCs) and T lymphocytes, thus affecting DCs activation, maturation, and priming of T cells involved in the immune response against intracellular pathogens. We studied a girl who developed at the age of 2.5 years aMycobacterium aviuminfection characterized by disseminated necrotizing granulomatous lymphadenitis, and we compared her findings with other patients with the same genetic condition. The patient carried a heterozygous 818del4 mutation in theIFNGR1gene responsible of autosomal dominant (AD) partial IFN gamma R1 deficiency. During the acute infection blood cells immunophenotyping showed a marked reduction in DCs counts, including both myeloid (mDCs) and plasmacytoid (pDCs) subsets, that reversed after successful prolonged antimicrobial therapy. Histology of her abdomen lymph node revealed a profound depletion of tissue pDCs, as compared to other age-matched granulomatous lymphadenitis of mycobacterial origin. Circulating DCs depletion was also observed in another patient with AD partial IFN gamma R1 deficiency during mycobacterial infection. To conclude, AD partial IFN gamma R1 deficiency can be associated with a transient decrease in both circulating and tissular DCs during acute mycobacterial infection, suggesting that DCs counts monitoring might constitute a useful marker of treatment response.