Substrate-induced DNA strand misalignment during catalytic cycling by DNA polymerase λ

The simple deletion of nucleotides is common in many organisms. It can be advantageous when it activates genes beneficial to microbial survival in adverse environments, and deleterious when it mutates genes relevant to survival, cancer or degenerative diseases. The classical idea is that simple deletions arise by strand slippage. A prime opportunity for slippage occurs during DNA synthesis, but it remains unclear how slippage is controlled during a polymerization cycle. Here, we report crystal structures and molecular dynamics simulations of mutant derivatives of DNA polymerase λ bound to a primer–template during strand slippage. Relative to the primer strand, the template strand is in multiple conformations, indicating intermediates on the pathway to deletion mutagenesis. Consistent with these intermediates, the mutant polymerases generate single-base deletions at high rates. The results indicate that dNTP-induced template strand repositioning during conformational rearrangements in the catalytic cycle is crucial to controlling the rate of strand slippage

From the Community to the Clinic: Building Community Health Worker-Inclusive Healthcare Teams

Multidisciplinary clinical teams with strong community linkages can engage with patient needs and address the social determinants of health. Community Health Workers (CHWs) have emerged as one such embodiment of this cultural shift in improving delivery and coordination of care to reach patient communities. However, misunderstandings of CHWs’ contributions have limited their uptake into clinical teams. In partnership with the Southwestern Area Health Education Center, this project investigated the context and perspectives of CHW engagement in Connecticut, focusing barriers and facilitators of CHW integration into clinical teams. Through experimentation, innovation, and mutual learning, integrated, CHW-inclusive healthcare teams can begin taking the necessary steps to bridge the divide between the community and the clinic. Commitment to service should be meaningfully considered in the CHW hiring process. Being altruistic, compassionate, nonjudgmental, and service-oriented are important attributes facilitating camaraderie and trust within healthcare teams, while also enhancing CHWs’ long-term commitments to organizations. Purposively seeking out these traits during the onboarding process can foster a team dynamic anchored by CHWs mutual commitment to serving and connecting with patients. Organizations should foster mutual understanding and respect for the varied roles CHWs play. Clearly defining roles and responsibilities and demonstrating the value that CHWs bring outside of the context of clinical care can improve collaboration, encourage skills-sharing, and promote an organizational climate of respect. Organizational decision-makers should increase the visibility of CHWs and include CHWs in conversations and meetings with other clinical team members where added value can be consistently demonstrated and where mutual learning and collaboration can meaningfully occur. Healthcare organizations should critically consider how to holistically support the CHW workforce. Organizational levers that promote retention, mutual learning, networking, and management of job stress amongst CHWs can improve their ability to function effectively and contribute to a diverse team culture. The fact that CHWs not only navigate the disparate worlds of the community and the clinic but also endure the stress of managing complex intra- and extra-organizational relationships should be recognized, valued, and appreciated by organizational leadership.https://elischolar.library.yale.edu/ysph_pbchrr/1049/thumbnail.jp

Current and Future Remote Sensing of Harmful Algal Blooms in the Chesapeake Bay to Support the Shellfish Industry

Harmful algal bloom (HAB) species in the Chesapeake Bay can negatively impact fish, shellfish, and human health via the production of toxins and the degradation of water quality. Due to the deleterious effects of HAB species on economically and environmentally important resources, such as oyster reef systems, Bay area resource managers are seeking ways to monitor HABs and water quality at large spatial and fine temporal scales. The use of satellite ocean color imagery has proven to be a beneficial tool for resource management in other locations around the world where high-biomass, nearly monospecific HABs occur. However, remotely monitoring HABs in the Chesapeake Bay is complicated by the presence of multiple, often co-occurring, species and optically complex waters. Here we present a summary of common marine and estuarine HAB species found in the Chesapeake Bay, Alexandrium monilatum, Karlodinium veneficum, Margalefidinium polykrikoides, and Prorocentrum minimum, that have been detected from space using multispectral data products from the Ocean and Land Colour Imager (OLCI) sensor on the Sentinel-3 satellites and identified based on in situ phytoplankton data and ecological associations. We review how future hyperspectral instruments will improve discrimination of potentially harmful species from other phytoplankton communities and present a framework in which satellite data products could aid Chesapeake Bay resource managers with monitoring water quality and protecting shellfish resources

Aptamer-based multiplexed proteomic technology for biomarker discovery

Interrogation of the human proteome in a highly multiplexed and efficient manner remains a coveted and challenging goal in biology. We present a new aptamer-based proteomic technology for biomarker discovery capable of simultaneously measuring thousands of proteins from small sample volumes (15 [mu]L of serum or plasma). Our current assay allows us to measure ~800 proteins with very low limits of detection (1 pM average), 7 logs of overall dynamic range, and 5% average coefficient of variation. This technology is enabled by a new generation of aptamers that contain chemically modified nucleotides, which greatly expand the physicochemical diversity of the large randomized nucleic acid libraries from which the aptamers are selected. Proteins in complex matrices such as plasma are measured with a process that transforms a signature of protein concentrations into a corresponding DNA aptamer concentration signature, which is then quantified with a DNA microarray. In essence, our assay takes advantage of the dual nature of aptamers as both folded binding entities with defined shapes and unique sequences recognizable by specific hybridization probes. To demonstrate the utility of our proteomics biomarker discovery technology, we applied it to a clinical study of chronic kidney disease (CKD). We identified two well known CKD biomarkers as well as an additional 58 potential CKD biomarkers. These results demonstrate the potential utility of our technology to discover unique protein signatures characteristic of various disease states. More generally, we describe a versatile and powerful tool that allows large-scale comparison of proteome profiles among discrete populations. This unbiased and highly multiplexed search engine will enable the discovery of novel biomarkers in a manner that is unencumbered by our incomplete knowledge of biology, thereby helping to advance the next generation of evidence-based medicine

Five views of a secret: does cognition change during middle adulthood?

This study examined five aspects of change (or stability) in cognitive abilities in middle adulthood across a 12-year period. Data come from the Interdisciplinary Study on Adult Development. The sample consisted of N = 346 adults (43.8 years on average, 48.6% female). In total, 11 cognitive tests were administered to assess fluid and crystallized intelligence, memory, and processing speed. In a first series of analyses, strong measurement invariance was established. Subsequently, structural stability, differential stability, stability of divergence, absolute stability, and the generality of changes were examined. Factor covariances were shown to be equal across time, implying structural stability. Stability coefficients were around .90 for fluid and crystallized intelligence, and speed, indicating high, yet not perfect differential stability. The coefficient for memory was .58. Only in processing speed the variance increased across time, indicating heterogeneity in interindividual development. Significant mean-level changes emerged, with an increase in crystallized intelligence and decline in the other three abilities. A number of correlations among changes in cognitive abilities were significant, implying that cognitive change

Factorial validity and measurement invariance across gender groups of the German version of the Interpersonal Reactivity Index

The Interpersonal Reactivity Index (IRI) is the most widely used measure of empathy, but its factorial validity has been questioned. The present research investigates the factorial validity of the German adaptation of the IRI, the "Saarbrücker Persönlichkeitsfragebogen SPF-IRI". Confirmatory Factor Analyses (CFA) and Exploratory Structural Equation Modeling (ESEM) were used to test the theoretically predicted four-factor model. Across two subsamples ESEM outperformed CFA. Substantial cross-loadings were evident in ESEM. Measurement invariance (MI) across gender groups was tested using ESEM in the combined sample. Strict MI (invariant factor loadings, intercepts, residuals) could be established, and variances and covariances were also equal. Differences for latent means were evident. Women scored higher on fantasy, empathic concern, and personal distress. No significant differences were found for perspective taking. Mean differences were due to real differences on latent variables and not a result of measurement bias. Results support the factorial validity of the German SPF-IRI. The heterogeneity of empathy and the unclear differentiation between cognitive and emotional aspects might be a source for the unclear differentiation of scales

Validation of a social cohesion theoretical framework: a multiple group SEM strategy

Social cohesion dates back to the end of the nineteenth century. Back then, society experienced epochal transformations, as are also happening nowadays. Whenever there are epochal changes, a social order (cohesion) matter arises. The paper provides a conceptual scheme of social cohesion identifying its constituent dimensions subdivided by three spheres (macro, meso, micro) and two perspectives (objective and subjective). The overarching aim is to test the validity of the operationalization of the social cohesion model provided. Firstly, we conducted an exploratory factor analysis introducing an approach implemented in Mplus named exploratory structural equation modeling that shows several useful characteristics. Afterward, through a structural equation modeling approach, we performed several confirmatory factor analyses adopting a multiple group SEM strategy in order to cross-validate the social cohesion model

Factor structure and measurement invariance across various demographic groups and over time for the phq-9 in primary care patients in spain

The Patient Health Questionnaire (PHQ-9) is a widely-used screening tool for depression in primary care settings. The purpose of the present study is to identify the factor structure of the PHQ-9 and to examine the measurement invariance of this instrument across different sociodemographic groups and over time in a sample of primary care patients in Spain. Data came from 836 primary care patients enrolled in a randomized controlled trial (PsicAP study) and a subsample of 218 patients who participated in a follow-up assessment at 3 months. Confirmatory factor analysis (CFA) was used to test one- and two-factor structures identified in previous studies. Analyses of multiple-group invariance were conducted to determine the extent to which the factor structure is comparable across various demo- graphic groups (i.e., gender, age, marital status, level of education, and employment situa- tion) and over time. Both one-factor and two-factor re-specified models met all the pre- established fit criteria. However, because the factors identified in the two-factor model were highly correlated (r = .86), the one-factor model was preferred for its parsimony. Multi-group CFA indicated measurement invariance across different demographic groups and across time. The present findings suggest that physicians in Spain can use the PHQ-9 to obtain a global score for depression severity in different demographic groups and to reliably monitor changes over time in the primary care setting

Pharmacological Evaluation of the Long-Term Effects of Xanomeline on the M1 Muscarinic Acetylcholine Receptor

Xanomeline is a unique agonist of muscarinic receptors that possesses functional selectivity at the M1 and M4 receptor subtypes. It also exhibits wash-resistant binding to and activation of the receptor. In the present work we investigated the consequences of this type of binding of xanomeline on the binding characteristics and function of the M1 muscarinic receptor. Pretreatment of CHO cells that stably express the M1 receptor for 1 hr with increasing concentrations of xanomeline followed by washing and waiting for an additional 23 hr in control culture media transformed xanomeline-induced inhibition of [3H]NMS binding from monophasic to biphasic. The high-affinity xanomeline binding site exhibited three orders of magnitude higher affinity than in the case of xanomeline added directly to the binding assay medium containing control cells. These effects were associated with a marked decrease in maximal radioligand binding and attenuation of agonist-induced increase in PI hydrolysis and were qualitatively similar to those caused by continuous incubation of cells with xanomeline for 24 hr. Attenuation of agonist-induced PI hydrolysis by persistently-bound xanomeline developed with a time course that parallels the return of receptor activation by prebound xanomeline towards basal levels. Additional data indicated that blockade of the receptor orthosteric site or the use of a non-functional receptor mutant reversed the long-term effects of xanomeline, but not its persistent binding at an allosteric site. Furthermore, the long-term effects of xanomeline on the receptor are mainly due to receptor down-regulation rather than internalization

High Resolution Detection and Analysis of CpG Dinucleotides Methylation Using MBD-Seq Technology

Methyl-CpG binding domain protein sequencing (MBD-seq) is widely used to survey DNA methylation patterns. However, the optimal experimental parameters for MBD-seq remain unclear and the data analysis remains challenging. In this study, we generated high depth MBD-seq data in MCF-7 cell and developed a bi-asymmetric-Laplace model (BALM) to perform data analysis. We found that optimal efficiency of MBD-seq experiments was achieved by sequencing ∼100 million unique mapped tags from a combination of 500 mM and 1000 mM salt concentration elution in MCF-7 cells. Clonal bisulfite sequencing results showed that the methylation status of each CpG dinucleotides in the tested regions was accurately detected with high resolution using the proposed model. These results demonstrated the combination of MBD-seq and BALM could serve as a useful tool to investigate DNA methylome due to its low cost, high specificity, efficiency and resolution