Infection of animal cells by numerous viruses is detected and countered by a variety of means, including recognition of nonself nucleic acids. The zinc finger antiviral protein (ZAP) depletes ...cytoplasmic RNA that is recognized as foreign in mammalian cells by virtue of its elevated CG dinucleotide content compared with endogenous mRNAs. Here, we determined a crystal structure of a protein-RNA complex containing the N-terminal, 4-zinc finger human (h) ZAP RNA-binding domain (RBD) and a CG dinucleotide-containing RNA target. The structure reveals in molecular detail how hZAP is able to bind selectively to CG-rich RNA. Specifically, the 4 zinc fingers create a basic patch on the hZAP RBD surface. The highly basic second zinc finger contains a pocket that selectively accommodates CG dinucleotide bases. Structure guided mutagenesis, cross-linking immunoprecipitation sequencing assays, and RNA affinity assays show that the structurally defined CG-binding pocket is not required for RNA binding per se in human cells. However, the pocket is a crucial determinant of high-affinity, specific binding to CG dinucleotide-containing RNA. Moreover, variations in RNA-binding specificity among a panel of CG-binding pocket mutants quantitatively predict their selective antiviral activity against a CG-enriched HIV-1 strain. Overall, the hZAP RBD RNA structure provides an atomic-level explanation for how ZAP selectively targets foreign, CG-rich RNA.
Using high-throughput screening we identified small molecules that suppress superoxide and/or H2O2 production during reverse electron transport through mitochondrial respiratory complex I (site IQ) ...without affecting oxidative phosphorylation (suppressors of site IQ electron leak, “S1QELs”). S1QELs diminished endogenous oxidative damage in primary astrocytes cultured at ambient or low oxygen tension, showing that site IQ is a normal contributor to mitochondrial superoxide-H2O2 production in cells. They diminished stem cell hyperplasia in Drosophila intestine in vivo and caspase activation in a cardiomyocyte cell model driven by endoplasmic reticulum stress, showing that superoxide-H2O2 production by site IQ is involved in cellular stress signaling. They protected against ischemia-reperfusion injury in perfused mouse heart, showing directly that superoxide-H2O2 production by site IQ is a major contributor to this pathology. S1QELs are tools for assessing the contribution of site IQ to cell physiology and pathology and have great potential as therapeutic leads.
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•S1QELs suppress site IQ ROS production without affecting oxidative phosphorylation•Site IQ generates superoxide-H2O2 in resting cells and in stress signaling•Site IQ drives ischemia-reperfusion injury in perfused heart and S1QELs protect•S1QELs are tools to assess the contribution of site IQ and therapeutic leads
In this article, Brand et al. identified small molecules (S1QELs) that suppress superoxide-H2O2 production during reverse electron transport through mitochondrial respiratory complex I (site IQ) without affecting oxidative phosphorylation. S1QELs diminished endogenous oxidative damage in primary astrocytes, stress signaling in vivo, and ischemia-reperfusion heart injury.
Mitochondrial electron transport drives ATP synthesis but also generates reactive oxygen species, which are both cellular signals and damaging oxidants. Superoxide production by respiratory complex ...III is implicated in diverse signaling events and pathologies, but its role remains controversial. Using high-throughput screening, we identified compounds that selectively eliminate superoxide production by complex III without altering oxidative phosphorylation; they modulate retrograde signaling including cellular responses to hypoxic and oxidative stress.
Pediatric nephrology is dedicated to caring for children with kidney disease, a unique blend of acute care and chronic longitudinal patient relationships. Though historically a small field, trainee ...interest has declined over the past 2 decades. This has led to growing alarm about the health of the pediatric nephrology workforce, although concerns have been hampered by a lack of available data to enable feasible projections. This article is part of a supplement that anticipates the future pediatric subspecialty workforce supply. It draws on existing literature, data from the American Board of Pediatrics, and findings from a model that estimates the future supply of pediatric subspecialists developed by the Carolina Health Workforce Research Center at the University of North Carolina Chapel Hill's Cecil G. Sheps Center for Health Services Research and Strategic Modeling Analytics & Planning Ltd. The workforce projections from 2020 to 2040 incorporate population growth, clinical effort, and geographic trends and model alternate scenarios adjusting for changes in trainee interest, clinical efforts, and workforce attrition. The baseline model predicts growth of clinical work equivalents by 26% by 2040, but further widening geographic disparities worsen the existing mismatch between supply, clinical need, and market demand. The worst-case scenario projects 13% growth by 2040 which, at best, maintains the status quo of an already strained workforce. The models do not account for many factors expected to heighten demand over the coming decades. Urgent reforms are necessary now. Proposed solutions require multipronged changes in education and training pathways, remuneration, clinical practice models, and government policy.
Zika virus (ZIKV) infection of pregnant women can cause fetal microcephaly and other neurologic defects. We describe the development of a non-human primate model to better understand fetal ...pathogenesis. To reliably induce fetal infection at defined times, four pregnant rhesus macaques are inoculated intravenously and intraamniotically with ZIKV at gestational day (GD) 41, 50, 64, or 90, corresponding to first and second trimester of gestation. The GD41-inoculated animal, experiencing fetal death 7 days later, has high virus levels in fetal and placental tissues, implicating ZIKV as cause of death. The other three fetuses are carried to near term and euthanized; while none display gross microcephaly, all show ZIKV RNA in many tissues, especially in the brain, which exhibits calcifications and reduced neural precursor cells. Given that this model consistently recapitulates neurologic defects of human congenital Zika syndrome, it is highly relevant to unravel determinants of fetal neuropathogenesis and to explore interventions.
The optimization for selectivity and central receptor occupancy for a series of spirocyclic azetidine–piperidine inverse agonists of the ghrelin receptor is described. Decreased mAChR muscarinic M2 ...binding was achieved by use of a chiral indane in place of a substituted benzylic group. Compounds with desirable balance of human in vitro clearance and ex vivo central receptor occupancy were discovered by incorporation of heterocycles. Specifically, heteroaryl rings with nitrogen(s) vicinal to the indane linkage provided the most attractive overall properties.
Using high-throughput screening we identified small molecules that suppress superoxide and/or H
O
production during reverse electron transport through mitochondrial respiratory complex I (site I
) ...without affecting oxidative phosphorylation (suppressors of site I
electron leak, "S1QELs"). S1QELs diminished endogenous oxidative damage in primary astrocytes cultured at ambient or low oxygen tension, showing that site I
is a normal contributor to mitochondrial superoxide-H
O
production in cells. They diminished stem cell hyperplasia in Drosophila intestine in vivo and caspase activation in a cardiomyocyte cell model driven by endoplasmic reticulum stress, showing that superoxide-H
O
production by site I
is involved in cellular stress signaling. They protected against ischemia-reperfusion injury in perfused mouse heart, showing directly that superoxide-H
O
production by site I
is a major contributor to this pathology. S1QELs are tools for assessing the contribution of site I
to cell physiology and pathology and have great potential as therapeutic leads.
Today's surface ocean is saturated with respect to calcium carbonate, but increasing atmospheric carbon dioxide concentrations are reducing ocean pH and carbonate ion concentrations, and thus the ...level of calcium carbonate saturation. Experimental evidence suggests that if these trends continue, key marine organisms--such as corals and some plankton--will have difficulty maintaining their external calcium carbonate skeletons. Here we use 13 models of the ocean-carbon cycle to assess calcium carbonate saturation under the IS92a 'business-as-usual' scenario for future emissions of anthropogenic carbon dioxide. In our projections, Southern Ocean surface waters will begin to become undersaturated with respect to aragonite, a metastable form of calcium carbonate, by the year 2050. By 2100, this undersaturation could extend throughout the entire Southern Ocean and into the subarctic Pacific Ocean. When live pteropods were exposed to our predicted level of undersaturation during a two-day shipboard experiment, their aragonite shells showed notable dissolution. Our findings indicate that conditions detrimental to high-latitude ecosystems could develop within decades, not centuries as suggested previously.
Common genetic variants contribute to the observed variation in breast cancer risk for BRCA2 mutation carriers; those known to date have all been found through population-based genome-wide ...association studies (GWAS). To comprehensively identify breast cancer risk modifying loci for BRCA2 mutation carriers, we conducted a deep replication of an ongoing GWAS discovery study. Using the ranked P-values of the breast cancer associations with the imputed genotype of 1.4 M SNPs, 19,029 SNPs were selected and designed for inclusion on a custom Illumina array that included a total of 211,155 SNPs as part of a multi-consortial project. DNA samples from 3,881 breast cancer affected and 4,330 unaffected BRCA2 mutation carriers from 47 studies belonging to the Consortium of Investigators of Modifiers of BRCA1/2 were genotyped and available for analysis. We replicated previously reported breast cancer susceptibility alleles in these BRCA2 mutation carriers and for several regions (including FGFR2, MAP3K1, CDKN2A/B, and PTHLH) identified SNPs that have stronger evidence of association than those previously published. We also identified a novel susceptibility allele at 6p24 that was inversely associated with risk in BRCA2 mutation carriers (rs9348512; per allele HR = 0.85, 95% CI 0.80-0.90, P = 3.9 × 10(-8)). This SNP was not associated with breast cancer risk either in the general population or in BRCA1 mutation carriers. The locus lies within a region containing TFAP2A, which encodes a transcriptional activation protein that interacts with several tumor suppressor genes. This report identifies the first breast cancer risk locus specific to a BRCA2 mutation background. This comprehensive update of novel and previously reported breast cancer susceptibility loci contributes to the establishment of a panel of SNPs that modify breast cancer risk in BRCA2 mutation carriers. This panel may have clinical utility for women with BRCA2 mutations weighing options for medical prevention of breast cancer.
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