Toll-like receptor (TLR) signaling plays a critical role in innate and adaptive immune responses and must be tightly controlled. TLR4 uses LPS binding protein, MD-2, and CD14 as accessories to ...respond to LPS. We therefore investigated the presence of an analagous soluble cofactor that might assist in the recruitment of CpG oligonucleotides (CpG-ODNs) to TLR9. We report the identification of granulin as an essential secreted cofactor that potentiates TLR9-driven responses to CpG-ODNs. Granulin, an unusual cysteine-rich protein, bound to CpG-ODNs and interacted with TLR9. Macrophages from granulin-deficient mice showed not only impaired delivery of CpG-ODNs to endolysosomal compartments, but also decreased interaction of TLR9 with CpG-ODNs. As a consequence, granulin-deficient macrophages showed reduced responses to stimulation with CpG-ODNs, a trait corrected by provision of exogenous granulin. Thus, we propose that granulin contributes to innate immunity as a critical soluble cofactor for TLR9 signaling.
► Granulin binds TLR9 and CpG oligonucleotides ► Granulin is sufficient for intracellular localization of CpG-ODNs ► Granulin is a critical cofactor in enabling TLR9 signal transduction
Regioregular head‐to‐tail (HT)‐coupled poly(3‐hexylthiophene‐2,5‐diyl) (P3HT) with a weight‐average molecular weight (Mw) in the 7.3–69.6 kDa range is crystallized by directional epitaxial ...solidification in 1,3,5‐trichlorobenzene (TCB) to yield highly oriented thin films. An oriented and periodic lamellar structure consisting of crystalline lamellae separated by amorphous interlamellar zones is evidenced by atomic force microscopy (AFM) and transmission electron microscopy (TEM). Both the overall crystallinity as well as the orientation of the crystalline lamellae decrease significantly with increasing Mw. The total lamellar periodicity is close to the length of “fully extended” chains for Mw = 7.3 kDa (polystyrene‐equivalent molecular weight, eq. PS) and it saturates to a value of ca. (25–28) ± 2 nm for Mw ≥ 18.8 kDa (eq. PS). This behavior is attributed to a transition from an oligomeric‐like system, for which P3HT chains are essentially in a fully extended all‐trans conformation and do not fold, to a semicrystalline system that involves a periodic alternation of crystalline lamellae separated by extended amorphous interlamellar zones, which harbor chain folds, chain ends, and tie molecules. For P3HT with Mw of ca. 7.3 kDa (eq. PS), epitaxial crystallization on TCB allows for the growth of both “edge‐on” and “flat‐on” oriented crystalline lamellae on the TCB substrate. The orientation of the lamellae is attributed to 1D epitaxy. Because of the large size of the “flat‐on” crystalline lamellae, a characteristic single‐crystal electron diffraction pattern corresponding to the 001 zone was obtained by selected area electron diffraction (SAED), indicating that P3HT crystallizes in a monoclinic unit cell with a = 16.0 Å, b = 7.8 Å, c = 7.8 Å, and γ = 93.5°.
Oriented thin films of poly(3‐hexylthiophene‐2,5‐diyl) (P3HT) with varying molecular weights are grown by directional epitaxial crystallization. P3HT with a weight‐average molecular weight Mw = 7.3 kDa behaves like an oligomeric system, whereas, for Mw ≥ 18.8 kDa, it shows the periodic lamellar structure of a semicrystalline polymer as a result of chain folding (see figure).
Toll-like receptor-9 (TLR9) is largely responsible for discriminating self from pathogenic DNA. However, association of host DNA with autoantibodies activates TLR9, inducing the pathogenic secretion ...of type I interferons (IFNs) from plasmacytoid dendritic cells (pDCs). Here, we found that in response to DNA-containing immune complexes (DNA-IC), but not to soluble ligands, IFN-α production depended upon the convergence of the phagocytic and autophagic pathways, a process called microtubule-associated protein 1A/1B-light chain 3 (LC3)-associated phagocytosis (LAP). LAP was required for TLR9 trafficking into a specialized interferon signaling compartment by a mechanism that involved autophagy-related proteins, but not the conventional autophagic preinitiation complex, or adaptor protein-3 (AP-3). Our findings unveil a new role for nonconventional autophagy in inflammation and provide one mechanism by which anti-DNA autoantibodies, such as those found in several autoimmune disorders, bypass the controls that normally restrict the apportionment of pathogenic DNA and TLR9 to the interferon signaling compartment.
► DNA-ICs recruit TLR9 and autophagy protein LC3 to phagosomes through FcγR signaling ► LAP, which is distinct from conventional autophagy, is required for IFN-α secretion ► LAP is not required for TLR9-mediated TNF secretion ► LAP allows trafficking of TLR9 into a specialized IFN-signaling compartment
A measurement and development platform for collecting water quality data (the WaterWatcher) was developed. The platform includes sensors to measure turbidity, total dissolved solids (TDS), and water ...temperature as variables that are often collected to assess water quality. The design is extensible for research and monitoring purposes, and all the design files are provided under open-source permissive licenses for further development. System design and operation are discussed for illustrative purposes. A block diagram indicates elements of mechanical, electrical, and software design for this system. The mechanical assembly used to house circuit boards and sensors is designed using 3D printing for rapid prototyping. The electronic circuit board acts as a carrier for an Arduino 32-bit microcontroller board and an associated cellular module along with a GPS for geolocation of water quality measurements. The cellular module permits data transfer for Internet of Things (IoT) functionality. System operation is set up using a command line interface (CLI) and C + + code that allows for calibration coefficients and human-readable transfer functions to be defined so that sensor voltages are related to physical quantities. Data are cached on a secure digital (SD) card for backup. The circuit was calibrated, and system operation assessed by deployment on an urban reservoir. Biogeochemical cycles were identified in the collected data using spectrogram and semivariogram analyses to validate system operation. As a system with hardware and software released under an open source license, the WaterWatcher platform reduces the time and effort required to build and deploy low-cost water quality measurement sensors and provides an example of the basic hardware design that can be used for measurements of water quality.
In the case of B.1.1.7, estimates suggest R could increase from 1 to about 1.4 with no change in population behavior.3,4 If true, many countries that have succeeded in reducing R to 1 or less will be ...confronted with a novel wave of viral spread despite the current measures.8,9 Once a more contagious variant has established itself, stabilising the number of new infections will become increasingly difficult. The joint action of all European countries will make each national and local effort more effective and impactful and safeguard public health across Europe.8 The longer restrictions last, and the less effective they become, the more depleted people's psychological, social, and economic resources become. Where novel variants require even stricter and longer measures than existing measures, it is of utmost importance to ensure that people with particularly heavy burdens receive financial and social support, that social burdens are justly distributed, and that mental health services meet the increasing demand to cope with bereavement, isolation, loss of income, fear, alcohol and drug misuse, insomnia, and anxiety as a result of the pandemic and lockdown strategies.
Overwhelming evidence shows that not only public health, but also society and the economy benefit greatly from reducing cases of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) ...infection. ...low case numbers save jobs and businesses. Making the case for the economic and social benefits of reducing case numbers will, if clearly communicated, greatly foster public cooperation.
We describe a multifactorial investigation of a SARS‐CoV‐2 outbreak in a large meat processing complex in Germany. Infection event timing, spatial, climate and ventilation conditions in the ...processing plant, sharing of living quarters and transport, and viral genome sequences were analyzed. Our results suggest that a single index case transmitted SARS‐CoV‐2 to co‐workers over distances of more than 8 m, within a confined work area in which air is constantly recirculated and cooled. Viral genome sequencing shows that all cases share a set of mutations representing a novel sub‐branch in the SARS‐CoV‐2 C20 clade. We identified the same set of mutations in samples collected in the time period between this initial infection cluster and a subsequent outbreak within the same factory, with the largest number of confirmed SARS‐CoV‐2 cases in a German meat processing facility reported so far. Our results indicate climate conditions, fresh air exchange rates, and airflow as factors that can promote efficient spread of SARS‐CoV‐2 via long distances and provide insights into possible requirements for pandemic mitigation strategies in industrial workplace settings.
Synopsis
There has been considerable debate about the factors contributing to SARS‐CoV‐2 outbreaks in food processing facilities around the world. This multifactorial investigation of an outbreak in a German meat processing plant shows that transmission occurred in a confined working area over long distances.
Analysis of infection event timing, spatial, climate and ventilation conditions, living quarters and transport, and viral genome sequences suggests a super spreading event that originated from a single employee.
Infections among workers over a distance of 8 m from the index case suggest aerosol transmission of SARS‐CoV-2.
The facilities’ environmental conditions such as low temperature, low air exchange rates, and constant air recirculation, together with relatively close distance between workers and demanding physical work, may have promoted efficient aerosol transmission.
In contrast to work‐related exposure, shared apartments, bedrooms, or carpools appear not to have played a major role in the initial outbreak.
Viral genome sequencing reveals a characteristic set of mutations that was also observed in samples collected during a later, much larger outbreak occurring in the same processing plant.
There has been considerable debate about the factors contributing to SARS‐CoV‐2 outbreaks in food processing facilities around the world. This multifactorial investigation of an outbreak in a German meat processing plant shows that transmission occurred in a confined working area over long distances.
Recent findings that 2-anilo-5-(4-methylpentan-2-yl)aminocyclohexa-2,5-diene-1,4-dione (6PPD-quinone), the transformation product of a common tire rubber antioxidant, is acutely toxic in ...stormwater-impacted streams has highlighted the need for a better understanding of contaminants in urban runoff. This study represents one of the first reports of 6PPD-quinone and other tire rubber-derived compounds in stormwater and snowmelt of a cold-climate Canadian city (Saskatoon, 2019–2020). Semiquantification of the five target compounds, N,N′-diphenylguanidine (DPG), N,N-dicyclohexylmethylamine (DCA), N,N′-dicyclohexylurea (DCU), 1-cyclohexyl-3-phenylurea (CPU), and 6PPD-quinone, revealed DPG was most abundant, with average concentrations of 60 μg L–1 in stormwater and 1 μg L–1 in snowmelt. Maximum observed concentrations of DPG were greater than 300 μg L–1, equivalent to loadings of 15 kg from a single rain event. These concentrations of DPG represent some of the highest reported in urban runoff globally. 6PPD-Quinone was detected in 57% (12/21) of stormwater samples with a mean concentration of approximately 600 ng L–1 (2019) and greater than 80% (28/31) of snowmelt samples with mean concentrations of 80–370 ng L–1 (2019 and 2020). Concentrations of 6PPD-quinone exceeded the acute LC50 for coho salmon (0.8–1.2 μg L–1) in greater than 20% of stormwater samples. Mass loadings of all target chemicals correlated well with roads and residential land-use area.
The latency-associated nuclear antigen (LANA) of Kaposi sarcoma herpesvirus (KSHV) is mainly localized and functions in the nucleus of latently infected cells, playing a pivotal role in the ...replication and maintenance of latent viral episomal DNA. In addition, N-terminally truncated cytoplasmic isoforms of LANA, resulting from internal translation initiation, have been reported, but their function is unknown. Using coimmunoprecipitation and MS, we found the cGMP-AMP synthase (cGAS), an innate immune DNA sensor, to be a cellular interaction partner of cytoplasmic LANA isoforms. By directly binding to cGAS, LANA, and particularly, a cytoplasmic isoform, inhibit the cGAS-STING–dependent phosphorylation of TBK1 and IRF3 and thereby antagonize the cGAS-mediated restriction of KSHV lytic replication. We hypothesize that cytoplasmic forms of LANA, whose expression increases during lytic replication, inhibit cGAS to promote the reactivation of the KSHV from latency. This observation points to a novel function of the cytoplasmic isoforms of LANA during lytic replication and extends the function of LANA from its role during latency to the lytic replication cycle.
Unc-93 homolog B1 (UNC93B1) is a key regulator of nucleic acid (NA)-sensing Toll-like receptors (TLRs). Loss of NA-sensing TLR responses in UNC93B1-deficient patients facilitates Herpes simplex virus ...type 1 (HSV-1) encephalitis. UNC93B1 is thought to guide NA-sensing TLRs from the endoplasmic reticulum (ER) to their respective endosomal signaling compartments and to guide the flagellin receptor TLR5 to the cell surface, raising the question of how UNC93B1 mediates differential TLR trafficking. Here, we report that UNC93B1 regulates a step upstream of the differential TLR trafficking process. We discovered that UNC93B1 deficiency resulted in near-complete loss of TLR3 and TLR7 proteins in primary splenic mouse dendritic cells and macrophages, showing that UNC93B1 is critical for maintaining TLR expression. Notably, expression of an ER-retained UNC93B1 version was sufficient to stabilize TLRs and largely restore endosomal TLR trafficking and activity. These data are critical for an understanding of how UNC93B1 can regulate the function of a broad subset of TLRs.
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•The trafficking of UNC93B1 and its client TLRs are separable events•ER-retained UNC93B1 is sufficient to restore UNC93B1-dependent TLR activities•UNC93B1 is essential for stabilizing nucleic-acid-sensing TLRs•UNC93B1-dependent TLR proteins are severely reduced in Unc93b1−/− and Unc93b13d/3d mice
UNC93B1 is known as a trafficking chaperone for both endosomal TLRs and cell-surface TLR5, but how UNC93B1 transports its client TLRs to different compartments remains to be determined. Pelka and colleagues show that, in fact, UNC93B1 functions upstream of differential trafficking pathways and is crucial for stabilizing UNC93B1-dependent TLRs.
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