The rockefeller university

In pathogenic mycobacteria, transcriptional responses to antibiotics result in induced antibiotic resistance. WhiB7 belongs to the Actinobacteria-specific family of Fe-S-containing transcription factors and plays a crucial role in inducible antibiotic resistance in mycobacteria. Here, we present cryoelectron microscopy structures of Mycobacterium tuberculosis transcriptional regulatory complexes comprising RNA polymerase sigma(A)-holoenzyme, global regulators CarD and RbpA, and WhiB7, bound to a WhiB7-regulated promoter. The structures reveal how WhiB7 interacts with sigma(A)-holoenzyme while simultaneously interacting with an AT-rich sequence element via its AT-hook. Evidently, AT-hooks, rare elements in bacteria yet prevalent in eukaryotes, bind to target AT-rich DNA sequences similarly to the nuclear chromosome binding proteins. Unexpectedly, a subset of particles contained a WhiB7-stabilized closed promoter complex, revealing this intermediate's structure, and we apply kinetic modeling and biochemical assays to rationalize how WhiB7 activates transcription. Altogether, our work presents a comprehensive view of how WhiB7 serves to activate gene expression leading to antibiotic resistance.

In Portugal, the 13-valent pneumococcal conjugate vaccine (PCV13) was commercially available between 2010 and 2015, following a decade of private use of PCV7. We evaluated changes on serotype distribution and antimicrobial susceptibility of pneumococci carried by children living in two regions of Portugal (one urban and one rural). Three epidemiological periods were defined: pre-PCV13 (2009-2010), early-PCV13 (2011-2012), and late-PCV13 (2015-2016). Nasopharyngeal samples (n = 4,232) were obtained from children 0-6 years old attending day-care centers. Private use of PCVs was very high in both regions (>75%). Pneumococcal carriage remained stable and high over time (62.1%, 62.4% and 61.6% (p = 0.909) in the urban region; and 59.8%, 62.8%, 59.5% (p = 0.543) in the rural region). Carriage of PCV7 serotypes remained low (5.3%, 7.8% and 4.3% in the urban region; and 2.5%, 3.7% and 4.8% in the rural region). Carriage of PCV13 serotypes not targeted by PCV7 decreased in both the urban (16.4%, 7.3%, and 1.6%; p < 0.001) and rural regions (13.2%, 7.8%, and 1.9%; p < 0.001). This decline was mostly attributable to serotype 19A (14.1%, 4.4% and 1.3% in the urban region; and 11.1%, 3.6% and 0.8% in the rural region, both p < 0.001). Serotype 3 declined over time in the urban region (10.1%, 4.4%, 0.8%; p < 0.001) and had no obvious trend in the rural region (4.2%, 6.7%, 2.4%; p = 0.505). Serotype 6C decreased in both regions while serotypes 11D, 15A/B/C, 16F, 21, 22F, 23A/B, 24F, 35F, and NT were the most prevalent in the late-PCV13 period. Intermediate resistance to penicillin and non-susceptibility to erythromycin decreased significantly in both regions (19.5%, 13.3%, and 9.3%; and 25.4%, 25.9%, and 13.4%; both p < 0.001, respectively in the urban region; and 12.4%, 11.1%, and 2.8% (p < 0.001); and 15.3%, 14.7%, and 9.2% (p = 0.037), respectively, in the rural region). In conclusion, private use of PCV13 led to significant changes on the pneumococcal population carried by children in Portugal. (c) 2021 The Authors. Published by Elsevier Ltd. This is an open access article under the CC BY license (http:// creativecommons.org/licenses/by/4.0/).

Human C2orf69 is an evolutionarily conserved gene whose function is unknown. Here, we report eight unrelated families from which 20 children presented with a fatal syndrome consisting of severe autoinflammation and progredient leukoencephalopathy with recurrent seizures; 12 of these subjects, whose DNA was available, segregated homozygous loss-of-function C2orf69 variants. C2ORF69 bears homology to esterase enzymes, and orthologs can be found in most eukaryotic genomes, including that of unicellular phytoplankton. We found that endogenous C2ORF69 (1) is loosely bound to mitochondria, (2) affects mitochondrial membrane potential and oxidative respiration in cultured neurons, and (3) controls the levels of the glycogen branching enzyme 1 (GBE1) consistent with a glycogen-storage-associated mitochondriopathy. We show that CRISPR-Cas9-mediated inactivation of zebrafish C2orf69 results in lethality by 8 months of age due to spontaneous epileptic seizures, which is preceded by persistent brain inflammation. Collectively, our results delineate an autoinflammatory Mendelian disorder of C2orf69 deficiency that disrupts the development/homeostasis of the immune and central nervous systems.

Listeria monocytogenes (L. monocytogenes) is a food-borne bacterial pathogen. Innate immunity to L. monocytogenes is profoundly affected by type I interferons (IFN-I). Here we investigated host metabolism in L. monocytogenes-infected mice and its potential control by IFN-I. Accordingly, we used animals lacking either the IFN-I receptor (IFNAR) or IRF9, a subunit of ISGF3, the master regulator of IFN-I-induced genes. Transcriptomes and metabolite profiles showed that L. monocytogenes infection induces metabolic rewiring of the liver. This affects various metabolic pathways including fatty acid (FA) metabolism and oxidative phosphorylation and is partially dependent on IFN-I signaling. Livers and macrophages from Ifnar1(-/-) mice employ increased glutaminolysis in an IRF9-independent manner, possibly to readjust TCA metabolite levels due to reduced FA oxidation. Moreover, FA oxidation inhibition provides protection from L. monocytogenes infection, explaining part of the protection of Irf9(-/-) and Ifnar1(-/-) mice. Our findings define a role of IFN-I in metabolic regulation during L. monocytogenes infection. Metabolic differences between Irf9(-/-) and Ifnar1(-/-) mice may underlie the different susceptibility of these mice against lethal infection with L. monocytogenes. Author summary Many immune cells undergo metabolic remodeling following encounters with cytokines or pathogenic insults. This is essential to perform their downstream effector functions and eradicate the infectious agents. Drug-mediated interference with metabolic remodeling can have a strong impact on clearance of the pathogen. Here we describe metabolic changes occurring during Listeria monocytogenes (L. monocytogenes) infection of murine hosts. Infected animals show profound changes of liver metabolism that include increased glycolysis, alterations in TCA cycle metabolites and the ratio of free versus conjugated fatty acids. Similar to what has been described during viral infections, type I interferons (IFN-I), a family of cytokines produced during L. monocytogenes infections, are involved in the import of fatty acids (FA) into the mitochondria to generate energy via oxidative phosphorylation. Accordingly, in the absence of IFN-I signals, the cells oxidize less FAs and this might help the cells better fight the infection. We also speculate that infected cells instead boost glutamine utilization to supply their energy need. Our work describes metabolic rewiring that takes place during L. monocytogenes infection and the contribution of IFN-I signaling. Our study improves the understanding of listeriosis and has the potential to help us discover new drug targets against L. monocytogenes and viruses that induce IFN-I response.

Germinal centers (GCs) are the site of immunoglobulin somatic hypermutation and affinity maturation, processes essential to an effective antibody response. The formation of GCs has been studied in detail, but less is known about what leads to their regression and eventual termination, factors that ultimately limit the extent to which antibodies mature within a single reaction. We show that contraction of immunization-induced GCs is immediately preceded by an acute surge in GC-resident Foxp3(+) T cells, attributed at least partly to up-regulation of the transcription factor Foxp3 by T follicular helper (T-FH) cells. Ectopic expression of Foxp3 in T-FH cells is sufficient to decrease GC size, implicating the natural up-regulation of Foxp3 by T-FH cells as a potential regulator of GC lifetimes.

The lymphatic vasculature plays important roles in the physiology of the organs in which it resides, though a clear mechanistic understanding of how this crosstalk is mediated is lacking. Here, we performed single-cell transcriptional profiling of human and mouse adipose tissue and found that lymphatic endothelial cells highly express neurotensin (NTS/Nts). Nts expression is reduced by cold and norepinephrine in an a-adrenergicdependent manner, suggesting a role in adipose thermogenesis. Indeed, NTS treatment of brown adipose tissue explants reduced expression of thermogenic genes. Furthermore, adenoviral-mediated overexpression and knockdown or knockout of NTS in vivo reduced and enhanced cold tolerance, respectively, an effect that is mediated by NTSR2 and ERK signaling. Inhibition of NTSR2 promoted energy expenditure and improved metabolic function in obese mice. These data establish a link between adipose tissue lymphatics and adipocytes with potential therapeutic implications.

Background: Large-scale epidemiological studies of seroprevalence of antibodies against SARS-CoV-2 often rely on point-of-care tests that provide immediate results to participants. Yet, little is known on how long rapid tests remain positive after the COVID-19 episode, or how much variability exists across different brands and even among batches of the same test. Methods: In November 2020, we assessed the sensitivity of three tests applied to 133 individuals with a previous positive PCR result between April and October. All subjects provided finger prick blood samples for two batches (A and B) of the Wondfo lateral -flow IgG/IgM test, and dried blood spot samples for the S-UFRJ ELISA test. Results: Overall sensitivity levels were 92.5% (95% CI 86.6-96.3), 63.2% (95% CI 54.4-71.4) and 33.8% (95% CI 25.9-42.5) for the S-UFRJ test, Wondfo A and Wondfo B tests, respectively. There was no evidence of a decline in the positivity of S-UFRJ with time since the diagnosis, but the two Wondfo batches showed sharp reductions to as low as 41.9% and 19.4%, respectively, for subjects with a positive PCR in June or earlier. Positive results for batch B of the rapid test were 35% to 54% lower than for batch A at any given month of diagnosis. Interpretation: Whereas the ELISA test showed high sensitivity and stability of results over the five months of the study, both batches of the rapid test showed substantial declines, with one of the batches consistently showing lower sensitivity levels than the other. ELISA tests based on dried-blood spots are an inexpensive alternative to rapid lateral-flow tests in large-scale epidemiological studies. Funding: The study was funded by the "Todos Pela Saude" initiative, Instituto Serrapilheira, Brazilian Ministry of Health, Brazilian Collective Health Association (ABRASCO) and the JBS S.A. initiative 'Fazer o Bem Faz Bem'. (C) 2021 Sociedade Brasileira de Infectologia. Published by Elsevier Espana, S.L.U.

Fanconi anemia is an inherited genome instability syndrome characterized by interstrand cross-link hypersensitivity, congenital defects, bone marrow failure, and cancer predisposition. Although DNA repair mediated by Fanconi anemia genes has been extensively studied, how inactivation of these genes leads to specific cellular phenotypic consequences associated with Fanconi anemia is not well understood. Here we report that Fanconi anemia stem cells in the C. elegans germline and in murine embryos display marked nonhomologous end joining (NHEJ)-dependent radiation resistance, leading to survival of progeny cells carrying genetic lesions. In contrast, DNA cross-linking does not induce generational genomic instability in Fanconi anemia stem cells, as widely accepted, but rather drives NHEJ-dependent apoptosis in both species. These findings suggest that Fanconi anemia is a stem cell disease reflecting inappropriate NHEJ, which is mutagenic and carcinogenic as a result of DNA misrepair, while marrow failure represents hematopoietic stem cell apoptosis. Significance: This study finds that Fanconi anemia stem cells preferentially activate error-prone NHEJ-dependent DNA repair to survive irradiation, thereby conferring generational genomic instability that is instrumental in carcinogenesis.

Improving clinical care for individuals infected with SARS-CoV-2 variants is a global health priority. Small-molecule antivirals like remdesivir (RDV) and biologics such as human monoclonal antibodies (mAbs) have demonstrated therapeutic efficacy against SARS-CoV-2, the causative agent of coronavirus disease 2019 (COVID-19). It is not known whether combination RDV/mAb will improve outcomes over single-agent therapies or whether antibody therapies will remain efficacious against variants. Here, we show that a combination of two mAbs in clinical trials, C144 and C135, have potent antiviral effects against even when initiated 48 h after infection and have therapeutic efficacy in vivo against the B.1.351 variant of concern (VOC). Combining RDV and antibodies provided a modest improvement in outcomes compared with single agents. These data support the continued use of RDV to treat SARS-CoV-2 infections and the continued clinical development of the C144 and C135 antibody combination to treat patients infected with SARS-CoV-2 variants.