Skovgaardbush4246

Transmission of COVID-19 from people without symptoms confounds societal mitigation strategies. From April to June 2020, we tested nasopharyngeal swabs by RT-qPCR from 15,514 staff and 16,966 residents of nursing homes and assisted living facilities in Massachusetts. Cycle threshold (Ct) distributions were very similar between populations with (N = 739) and without (N = 2179) symptoms at the time of sampling (mean Ct 25.7 versus 26.4, ranges 12-38). However, as local cases waned, those without symptoms shifted towards higher Ct. With such similar viral load distributions, existing testing modalities should perform comparably regardless of symptoms, contingent upon time since infection.T7 RNA polymerase (RNAP) is a valuable tool in biotechnology, basic research and synthetic biology due to its robust, efficient and selective transcription of genes. Here, we expand the scope of T7 RNAP to include plasmid replication. We present a novel type of plasmid, termed T7 ori plasmids that replicate, in an engineered Escherichia coli, with a T7 phage origin as the sole origin of replication. We find that while the T7 replication proteins; T7 DNA polymerase, T7 single-stranded binding proteins and T7 helicase-primase are dispensable for replication, T7 RNAP is required, although dependent on a T7 RNAP variant with reduced activity. We also find that T7 RNAP-dependent replication of T7 ori plasmids requires the inactivation of cellular ribonuclease H. We show that the system is portable among different plasmid architectures and ribonuclease H-inactivated E. coli strains. Finally, we find that the copy number of T7 ori plasmids can be tuned based on the induction level of RNAP. Altogether, this study assists in the choice of an optimal genetic tool by providing a novel plasmid that requires T7 RNAP for replication.Shelterin is a six-protein complex that coats chromosome ends to ensure their proper protection and maintenance. Similar to the human shelterin, fission yeast shelterin is composed of telomeric double- and single-stranded DNA-binding proteins, Taz1 and Pot1, respectively, bridged by Rap1, Poz1 and Tpz1. The assembly of the proteinaceous Tpz1-Poz1-Rap1 complex occurs cooperatively and disruption of this shelterin bridge leads to unregulated telomere elongation. However, how this biophysical property of bridge assembly is integrated into shelterin function is not known. Here, utilizing synthetic bridges with a range of binding properties, we find that synthetic shelterin bridge lacking cooperativity requires a linker pair that matches the native bridge in complex lifespan but has dramatically higher affinity. We find that cooperative assembly confers kinetic properties on the shelterin bridge allowing disassembly to function as a molecular timer, regulating the duration of the telomere open state, and consequently telomere lengthening to achieve a defined species-specific length range.Bacteria have evolved sophisticated mechanisms to deliver potent toxins into bacterial competitors or into eukaryotic cells in order to destroy rivals and gain access to a specific niche or to hijack essential metabolic or signaling pathways in the host. Delivered effectors carry various activities such as nucleases, phospholipases, peptidoglycan hydrolases, enzymes that deplete the pools of NADH or ATP, compromise the cell division machinery, or the host cell cytoskeleton. Effectors categorized in the family of polymorphic toxins have a modular structure, in which the toxin domain is fused to additional elements acting as cargo to adapt the effector to a specific secretion machinery. Here we show that Photorhabdus laumondii, an entomopathogen species, delivers a polymorphic antibacterial toxin via a type VI secretion system. This toxin inhibits protein synthesis in a NAD+-dependent manner. Using a biotinylated derivative of NAD, we demonstrate that translation is inhibited through ADP-ribosylation of the ribosomal 23S RNA. Mapping of the modification further showed that the adduct locates on helix 44 of the thiostrepton loop located in the GTPase-associated center and decreases the GTPase activity of the EF-G elongation factor.Translation of eukaryotic mRNAs begins with binding of their m7G cap to eIF4E, followed by recruitment of other translation initiation factor proteins. We describe capCLIP, a novel method to comprehensively capture and quantify the eIF4E (eukaryotic initiation factor 4E) 'cap-ome' and apply it to examine the biological consequences of eIF4E-cap binding in distinct cellular contexts. First, we use capCLIP to identify the eIF4E cap-omes in human cells with/without the mTORC1 (mechanistic target of rapamycin, complex 1) inhibitor rapamycin, there being an emerging consensus that rapamycin inhibits translation of TOP (terminal oligopyrimidine) mRNAs by displacing eIF4E from their caps. capCLIP reveals that the representation of TOP mRNAs in the cap-ome is indeed systematically reduced by rapamycin, thus validating our new methodology. capCLIP also refines the requirements for a functional TOP sequence. Second, we apply capCLIP to probe the consequences of phosphorylation of eIF4E. We show eIF4E phosphorylation reduces overall eIF4E-mRNA association and, strikingly, causes preferential dissociation of mRNAs with short 5'-UTRs. capCLIP is a valuable new tool to probe the function of eIF4E and of other cap-binding proteins such as eIF4E2/eIF4E3.Although autocatalytic ethylene biosynthesis plays an important role in the ripening of climacteric fruits, our knowledge of the network that promotes autocatalytic ethylene biosynthesis remains limited. We identified white fruit (wf), a tomato mutant that produces immature fruit that are white and that ripen slowly. We found that an inversion on chromosome 10 that disrupts the LUTESCENT2 gene, and the white fruit is allelic to lutescent 2. Using CRISPR-Cas9 technology we knocked out L2 in wild type tomato and found that the l2-cr mutants produced phenotype that were very similar to white fruit (lutescent 2). In the l2-cr fruit, chloroplast development was impaired and the accumulation of carotenoids and lycopene occurred more slowly than in wild type. During fruit ripening in l2-cr mutants, the peak of ethylene release was delayed, less ethylene was produced and the expression of ACO genes was significantly suppressed. We also found that exogenous ethylene induces the expression of L2 and that ERF.B3, an ethylene response factor, binds the promoter of the L2 gene and activates its transcription. Thus, the expression of L2 is regulated by exogenous ethylene. Taken together, our results indicate that ethylene may affect the expression of the L2 gene and that the L2 gene participates in autocatalytic ethylene biosynthesis during tomato fruit ripening.In the cell, stalled ribosomes are rescued through ribosome-associated protein quality-control (RQC) pathways. After splitting of the stalled ribosome, a C-terminal polyalanine 'tail' is added to the unfinished polypeptide attached to the tRNA on the 50S ribosomal subunit. In Bacillus subtilis, polyalanine tailing is catalyzed by the NEMF family protein RqcH, in cooperation with RqcP. However, the mechanistic details of this process remain unclear. Here we demonstrate that RqcH is responsible for tRNAAla selection during RQC elongation, whereas RqcP lacks any tRNA specificity. The ribosomal protein uL11 is crucial for RqcH, but not RqcP, recruitment to the 50S subunit, and B. subtilis lacking uL11 are RQC-deficient. Through mutational mapping, we identify critical residues within RqcH and RqcP that are important for interaction with the P-site tRNA and/or the 50S subunit. Additionally, we have reconstituted polyalanine-tailing in vitro and can demonstrate that RqcH and RqcP are necessary and sufficient for processivity in a minimal system. Moreover, the in vitro reconstituted system recapitulates our in vivo findings by reproducing the importance of conserved residues of RqcH and RqcP for functionality. Collectively, our findings provide mechanistic insight into the role of RqcH and RqcP in the bacterial RQC pathway.

Evaluating age as a risk factor for susceptibility to infectious diseases, particularly for COVID-19, is critical. Cytomegalovirus (CMV) serologic prevalence increases with age, and associates with inflammatory-mediated diseases in the elderly. However, little is known regarding the subclinical impact of CMV and risk it poses to healthy older adults. Prior to the COVID-19 pandemic we conducted a study to determine the association of CMV to biologic age and immune dysregulation.

Community-dwelling, healthy adults over 60 years old were evaluated using DNA methylation assays to define epigenetic age (EpiAge) and T cell immunophenotyping to assess immune dysregulation.

All subjects were healthy and asymptomatic. Those CMV seropositive had more lymphocytes, CD8 T cells, CD28 negative T cells, decreased CD4/CD8 cell ratios, and had higher average EpiAge (65.34 years) than those CMV seronegative (59.53 years). Decreased % CD4 (p=0.003) and numbers of CD4 T cells (p=0.0199) correlated to increased EpiAge.

Our novel findings distinguish altered immunity in the elderly based on CMV status. Chronic CMV infection in healthy, older adults is associated with indicators of immune dysregulation, both of which correlate to differences in EpiAge.

Our novel findings distinguish altered immunity in the elderly based on CMV status. Chronic CMV infection in healthy, older adults is associated with indicators of immune dysregulation, both of which correlate to differences in EpiAge.Recent technical advance attracts great attention to the promotion of programming skills, in particular, and computational thinking (CT), in general, as a new intellectual competency. However, the understanding of its cognitive substrates is limited. The present study used functional magnetic resonance imaging to examine the neural correlates of programming to understand the cognitive substrates of CT. Specifically, magnetic resonance imaging signals were collected while the participants were mentally solving programming problems, and we found that CT recruited distributed cortical regions, including the posterior parietal cortex, the medial frontal cortex, and the left lateral frontal cortex. These regions showed extensive univariate and multivariate resemblance with arithmetic, reasoning, and spatial cognition tasks. Based on the resemblance, clustering analyses revealed that cortical regions involved in CT can be divided into Reasoning, Calculation, Visuospatial, and Shared components. Further, connectivity increased during programming within the CT network constructed by these four components and decreased between the CT network and other cortical regions. selleck inhibitor In sum, our study revealed the cognitive components underlying CT and their neural correlates and further suggests that CT is not a simple sum of parallel cognitive processes, but a composite cognitive process integrating a set of intellectual abilities, particularly those in the science, technology, engineering, and math domains.

tRNAs were originally considered uni-functional RNA molecules involved in the delivery of amino acids to growing peptide chains on the ribosome. More recently, the liberation of tRNA fragments from tRNAs via specific enzyme cleavage has been characterized. Detection of tRNA fragments in sequencing data is difficult due to tRNA sequence redundancy and the short length of both tRNAs and their fragments.

Here we introduce tsRNAsearch, a Nextflow pipeline for the identification of differentially abundant tRNA fragments and other non-coding RNAs from small RNA-sequencing data. tsRNAsearch is intended for use when comparing two groups of datasets, such as control and treatment groups. tsRNAsearch comparatively searches for tRNAs and ncRNAs with irregular read distribution profiles (a proxy for RNA cleavage) using a combined score made up of four novel methods and a differential expression analysis, and reports the top ranked results in simple PDF and TEXT files. In this study, we used publicly available small RNA-seq data to replicate the identification of tsRNAs from chronic hepatitis-infected liver tissue data.