The transcription factor NF-κB orchestrates nucleosome remodeling during the primary response to Toll-like receptor 4 signaling.

Inducible nucleosome remodeling at hundreds of latent enhancers and several promoters shapes the transcriptional response to Toll-like receptor 4 (TLR4) signaling in macrophages. We aimed to define the identities of the transcription factors that promote TLR-induced remodeling. An analysis strategy based on ATAC-seq and single-cell ATAC-seq that enriched for genomic regions most likely to undergo remodeling revealed that the transcription factor nuclear factor κB (NF-κB) bound to all high-confidence peaks marking remodeling during the primary response to the TLR4 ligand, lipid A. Deletion of NF-κB subunits RelA and c-Rel resulted in the loss of remodeling at high-confidence ATAC-seq peaks, and CRISPR-Cas9 mutagenesis of NF-κB-binding motifs impaired remodeling. Remodeling selectivity at defined regions was conferred by collaboration with other inducible factors, including IRF3- and MAP-kinase-induced factors. Thus, NF-κB is unique among TLR4-activated transcription factors in its broad contribution to inducible nucleosome remodeling, alongside its ability to activate poised enhancers and promoters assembled into open chromatin.

The psychophysics of affordance perception: Stevens' power law scaling of perceived maximum forward reachability with an object.

The literatures on affordance perception and psychophysics are seminal in the basic study of perception and action. Nevertheless, the application of classic psychophysical methodologies/analysis to the study of affordance perception remains unexplored. In four experiments, we investigated the Stevens' power law scaling of affordance perception. Participants reported maximum forward reaching ability with a series of rods (both seated and standing) for themselves and another person (confederate). Participants also reported a property of the rod set that has been explored in previous psychophysical experiments and changes in equal measure with forward reach-with-ability (length). In all, we found that affordance perception reports (ß = .32) were an underaccelerated function of actual changes in reaching ability compared with relatively less accelerated length reports (ß = .73). Affordance perception scaled with stimulus magnitude more similarly to brightness perception than length perception. Furthermore, affordance perception reports scaled similarly regardless of the actor (self and other), task context (seated and standing), or idiosyncrasies of the measurement procedure (controlling for distance compression effects), while length perception reports were sensitive to location/distance compression effects. We offer empirical and theoretical considerations, along with pathways for future research.

Detection of viral gene expression in risk-stratified biopsies reveals no active HPV in cutaneous squamous cell carcinoma.

BACKGROUND: Human papillomavirus (HPV) infection is known to promote the development of mucosal squamous cell carcinoma (mSCC), including pathologically high-grade lesions, but its role in cutaneous squamous cell carcinoma (cuSCC) remains unclear, particularly in lesions that are considered high risk. OBJECTIVE: We aimed to determine whether enhanced HPV transcriptional activity can be detected in high-risk cuSCC samples compared with low-grade SCC samples or normal skin. METHODS: We performed RNA sequencing of cuSCC across 23 risk-stratified skin lesions. A subset of samples was tested for the presence of HPV DNA. High-quality, non-human reads from each sample group were used for viral analysis using Microbiome Coverage Profiler. RESULTS: None of the samples analysed had detectable expression of HPV RNA, while 64% of samples tested positive for HPV DNA. All samples were found to have expression of human endogenous retrovirus, and multiple samples showed expression of other viruses. CONCLUSIONS: Viral and prophage gene expression can be monitored in cuSCC or normal skin biopsies, yet no sample in our study showed evidence of active HPV gene expression despite evidence of HPV genome presence. This suggests HPV transcription does not play a role in differentiating high-risk cuSCCs from low-risk cuSCCs or normal skin.

Analytical method development for characterizing ingredient-specific particle size distributions of nasal spray suspension products.

Particle size characterization for active pharmaceutical ingredients (APIs) in nasal spray suspension products presents unique challenges because both the API and excipient particles are present in the final dosage form. Currently, an established method is lacking because traditional particle sizing technologies do not distinguish the chemical identity of the particles. In this study, a non-destructive, ingredient-specific particle sizing method was developed for characterization of mometasone furoate (MF) nasal spray suspensions using Morphology Directed Raman Spectroscopy (MDRS). A five-step method development procedure was used in this study: sample preparation, particle imaging and morphology analysis, particle Raman measurements and classification, morphology filter selection, and minimum number of particles determination. Wet dispersion sample preparation method was selected to ensure that the particles were measured in their original suspended state. A training set containing over 10,000 randomly-selected particles, including both the API and excipient particles, was used to gain a comprehensive understanding of particle size, shape, and chemical ID for the nasal spray suspension. Morphology and Raman measurements were performed on each particle in the training set. The measurement results suggested that the aspect ratio and intensity mean filter combination was an appropriate morphology filter setting to selectively target API particles and exclude most of excipient particles. With further optimization of the morphology filter cutoff values and determination of minimal number of particles to be measured, the total measurement time was reduced from 90 hours to 8 hours. The morphologically screening strategy ultimately allowed us to create a time-efficient practical API-specific particle size distribution (PSD) methods for nasal spray suspensions. This study shows that MDRS is a fit for purpose analytical technique for determining ingredient-specific PSDs of the pharmaceutical formulation studied in this work.

Single cell analysis reveals immune cell-adipocyte crosstalk regulating the transcription of thermogenic adipocytes.

Immune cells are vital constituents of the adipose microenvironment that influence both local and systemic lipid metabolism. Mice lacking IL10 have enhanced thermogenesis, but the roles of specific cell types in the metabolic response to IL10 remain to be defined. We demonstrate here that selective loss of IL10 receptor α in adipocytes recapitulates the beneficial effects of global IL10 deletion, and that local crosstalk between IL10-producing immune cells and adipocytes is a determinant of thermogenesis and systemic energy balance. Single Nuclei Adipocyte RNA-sequencing (SNAP-seq) of subcutaneous adipose tissue defined a metabolically-active mature adipocyte subtype characterized by robust expression of genes involved in thermogenesis whose transcriptome was selectively responsive to IL10Rα deletion. Furthermore, single-cell transcriptomic analysis of adipose stromal populations identified lymphocytes as a key source of IL10 production in response to thermogenic stimuli. These findings implicate adaptive immune cell-adipocyte communication in the maintenance of adipose subtype identity and function.

Anxiety Influences the Perceptual-Motor Calibration of Visually Guided Braking to Avoid Collisions.

We investigated whether anxiety influences perceptual-motor calibration in a braking to avoid a collision task. Participants performed either a discrete braking task (Experiment 1) or a continuous braking task (Experiment 2), with the goal of stopping before colliding with a stop sign. Half of participants performed the braking task after an anxiety induction. We investigated whether anxiety reduced the frequency of crashing and if it influenced the calibration of perception (visual information) and action (brake pressure) dynamically between-trials in Experiment 1 and within-trials in Experiment 2. In the discrete braking task, anxious participants crashed less often and made larger corrective adjustments trial-to-trial after crashing, suggesting that the influence of anxiety on behavior did not occur uniformly, but rather dynamically with anxiety amplifying the reaction to previous crashes. However, when performing continuous braking, anxious participants crashed more often, and their within-trial adjustments of deceleration were less related to visual information compared to controls. Taken together, these findings suggest that the timescale and nature of the task mediates the influence of anxiety on the performance of goal-directed actions.

Not all those who wander are lost: Spatial exploration patterns and their relationship to gender and spatial memory.

When humans and animals navigate through environments, they form spatial memories important for supporting subsequent recall of locations relative to their own position and orientation, as well as to other object locations in the environment. The goal of the current study was to examine whether individual differences in initial exploration of a large-scale novel environment relate to subsequent spatial memories. A majority of studies examining spatial memory formed in large-scale spaces have constrained encoding of the environment by leading participants on pre-determined paths, thereby limiting their free exploration. We allowed participants to freely explore a large-scale, virtual environment to locate a set of objects within. We then tested their ability to navigate back to those objects as well as their ability to point to them from one another. Based on previous work suggesting gender differences in navigation strategies and spatial anxiety, we predicted that males and females would show different patterns of initial exploration and that these exploration patterns would account for gender differences in measures of spatial memory. We found that females revisited previous locations more often and showed lower rates of spreading through an area. These measures of exploration partially accounted for gender differences in efficiency in navigation and pointing accuracy to remembered locations. The results demonstrate the importance of exploration in spatial memory and provide a new perspective on gender differences in spatial cognition.

Transcriptional regulation of macrophage cholesterol efflux and atherogenesis by a long noncoding RNA.

Nuclear receptors regulate gene expression in response to environmental cues, but the molecular events governing the cell type specificity of nuclear receptors remain poorly understood. Here we outline a role for a long noncoding RNA (lncRNA) in modulating the cell type-specific actions of liver X receptors (LXRs), sterol-activated nuclear receptors that regulate the expression of genes involved in cholesterol homeostasis and that have been causally linked to the pathogenesis of atherosclerosis. We identify the lncRNA MeXis as an amplifier of LXR-dependent transcription of the gene Abca1, which is critical for regulation of cholesterol efflux. Mice lacking the MeXis gene show reduced Abca1 expression in a tissue-selective manner. Furthermore, loss of MeXis in mouse bone marrow cells alters chromosome architecture at the Abca1 locus, impairs cellular responses to cholesterol overload, and accelerates the development of atherosclerosis. Mechanistic studies reveal that MeXis interacts with and guides promoter binding of the transcriptional coactivator DDX17. The identification of MeXis as a lncRNA modulator of LXR-dependent gene expression expands understanding of the mechanisms underlying cell type-selective actions of nuclear receptors in physiology and disease.

IL-10 Signaling Remodels Adipose Chromatin Architecture to Limit Thermogenesis and Energy Expenditure.

Signaling pathways that promote adipose tissue thermogenesis are well characterized, but the limiters of energy expenditure are largely unknown. Here, we show that ablation of the anti-inflammatory cytokine IL-10 improves insulin sensitivity, protects against diet-induced obesity, and elicits the browning of white adipose tissue. Mechanistic studies define bone marrow cells as the source of the IL-10 signal and adipocytes as the target cell type mediating these effects. IL-10 receptor alpha is highly enriched in mature adipocytes and is induced in response to differentiation, obesity, and aging. Assay for transposase-accessible chromatin sequencing (ATAC-seq), ChIP-seq, and RNA-seq reveal that IL-10 represses the transcription of thermogenic genes in adipocytes by altering chromatin accessibility and inhibiting ATF and C/EBPß recruitment to key enhancer regions. These findings expand our understanding of the relationship between inflammatory signaling pathways and adipose tissue function and provide insight into the physiological control of thermogenesis that could inform future therapy.

Perceiver as polar planimeter: Direct perception of jumping, reaching, and jump-reaching affordances for the self and others.

Runeson (Scandanavian Journal of Psychology 18:172-179, 1977) suggested that the polar planimeter might serve as an informative model system of perceptual mechanism. The key aspect of the polar planimeter is that it registers a higher order property of the environment without computational mediation on the basis of lower order properties, detecting task-specific information only. This aspect was posited as a hypothesis for the perception of jumping and reaching affordances for the self and another person. The findings supported this hypothesis. The perception of reaching while jumping significantly differed from an additive combination of jump-without-reaching and reach-without-jumping perception. The results are consistent with Gibson's (The senses considered as perceptual systems, Houghton Mifflin, Boston, MA; Gibson, The senses considered as perceptual systems, Houghton Mifflin, Boston, MA, 1966; The ecological approach to visual perception, Houghton Mifflin, Boston, MA; Gibson, The ecological approach to visual perception, Houghton Mifflin, Boston, MA, 1979) theory of information-that aspects of the environment are specified by patterns in energetic media.

Perceiving and Remembering Affordances for Others Are Continuous Processes.

In information-based approaches, affordances are perceived by detecting information that specifies an animal-environment fit, not by combining perceptions of constituent lower-order properties. Given that detection of such information necessarily occurs over space and time, there is no clear distinction between perception and memory. Rather, perceiving and remembering are continuous processes. Whereas previous research has investigated the continuity of perceived and remembered affordances for the self, we did so with respect to perceived and remembered affordances for others. Participants reported remembered maximum reaching height and remembered anthropometric properties of another person. Remembered maximum reaching height was not reducible to a combination of remembered anthropometric properties. Moreover, remembered maximum reaching height scaled to the reaching ability of the other person and not to that of the perceiver. Both results are consistent with an information-based perspective on perceiving and remembering affordances and demonstrate a continuity between perceiving and remembering affordances for others.

Opposing roles of Toll-like receptor and cytosolic DNA-STING signaling pathways for Staphylococcus aureus cutaneous host defense.

Successful host defense against pathogens requires innate immune recognition of the correct pathogen associated molecular patterns (PAMPs) by pathogen recognition receptors (PRRs) to trigger the appropriate gene program tailored to the pathogen. While many PRR pathways contribute to the innate immune response to specific pathogens, the relative importance of each pathway for the complete transcriptional program elicited has not been examined in detail. Herein, we used RNA-sequencing with wildtype and mutant macrophages to delineate the innate immune pathways contributing to the early transcriptional response to Staphylococcus aureus, a ubiquitous microorganism that can activate a wide variety of PRRs. Unexpectedly, two PRR pathways-the Toll-like receptor (TLR) and Stimulator of Interferon Gene (STING) pathways-were identified as dominant regulators of approximately 95% of the genes that were potently induced within the first four hours of macrophage infection with live S. aureus. TLR signaling predominantly activated a pro-inflammatory program while STING signaling activated an antiviral/type I interferon response with live but not killed S. aureus. This STING response was largely dependent on the cytosolic DNA sensor cyclic guanosine-adenosine synthase (cGAS). Using a cutaneous infection model, we found that the TLR and STING pathways played opposite roles in host defense to S. aureus. TLR signaling was required for host defense, with its absence reducing interleukin (IL)-1ß production and neutrophil recruitment, resulting in increased bacterial growth. In contrast, absence of STING signaling had the opposite effect, enhancing the ability to restrict the infection. These results provide novel insights into the complex interplay of innate immune signaling pathways triggered by S. aureus and uncover opposing roles of TLR and STING in cutaneous host defense to S. aureus.

Note: A simple detection method for helium droplet spectroscopy experiments.

Helium droplet methods are currently established as a premier experimental technique for the production and spectroscopic study of novel clusters and complexes. Unfortunately, some of the essential equipment required to perform the experiments, such as the detector used to monitor photon-induced depletion of the helium droplet beam, can be relatively large, complex, and expensive. Most often this detector is a quadrupole mass spectrometer (QMS). In this report, we describe the development and evaluation of an extremely simple, straightforward, small, and inexpensive droplet beam detector for use in helium droplet spectroscopy experiments and compare its performance to that of a QMS by recording the infrared spectra of helium droplets doped with either 13CO2 or CD4.

The Independent Perceptual Calibration of Action-Neutral and -Referential Environmental Properties.

Two experiments were conducted to explore how the calibration of perception of environmental properties taken with reference to an animal and their action capabilities (e.g., affordances) and those that are independent of action capabilities (e.g., metric properties) relate. In both experiments, participants provided reports of the maximum height they could reach above their head with a number of different stick(s) (reach-with-stick height) and the length of those stick(s), a property that is a constituent of reach-with-stick height. In Experiment 1 reach-with-stick height reports improved over trials whereas stick length reports remained constant. In Experiment 2, feedback about maximum reach-with-stick height improved perception of this affordance, but such improvements did not transfer to perception of stick length in a pretest/practice task/posttest design. The results suggest that the perceptual calibration with practice perceiving or feedback about actual dimensions of action-referential and action-neutral properties do not necessarily depend on one another.

The Ly6C(lo)-Down: Targeting Developmental Enhancers for Cell Subset Depletion.

The diversity of mononuclear phagocytes has made it difficult to ascribe cellular functions to sub-populations using conventional loss-of-function approaches. In this issue of Immunity, Thomas et al. (2016) highlight the value of excising enhancer domains, effectively depleting defined lineages without altering other cellular physiology.

A Stringent Systems Approach Uncovers Gene-Specific Mechanisms Regulating Inflammation.

Much has been learned about transcriptional cascades and networks from large-scale systems analyses of high-throughput datasets. However, analysis methods that optimize statistical power through simultaneous evaluation of thousands of ChIP-seq peaks or differentially expressed genes possess substantial limitations in their ability to uncover mechanistic principles of transcriptional control. By examining nascent transcript RNA-seq, ChIP-seq, and binding motif datasets from lipid A-stimulated macrophages with increased attention to the quantitative distribution of signals, we identified unexpected relationships between the in vivo binding properties of inducible transcription factors, motif strength, and transcription. Furthermore, rather than emphasizing common features of large clusters of co-regulated genes, our results highlight the extent to which unique mechanisms regulate individual genes with key biological functions. Our findings demonstrate the mechanistic value of stringent interrogation of well-defined sets of genes as a complement to broader systems analyses of transcriptional cascades and networks.

The effect of semantic context on prospective memory performance.

The current study provides evidence for spontaneous processing in prospective memory (PM) or memory for intentions. Discrepancy-plus-search is the spontaneous processing of PM cues via disruptions in processing fluency of ongoing task items. We tested whether this mechanism can be demonstrated in an ongoing rating task with a dominant semantic context. Ongoing task items were manipulated such that the PM cues were members of a semantic category (i.e., Body Parts) that was congruent or discrepant with the dominant semantic category in the ongoing task. Results showed that participants correctly responded to more PM cues when there was a category discrepancy between the PM cues and ongoing task items. Moreover, participants' identification of PM cues was accompanied by faster ongoing task reaction times when PM cues were discrepant with ongoing task items than when they were congruent. These results suggest that a discrepancy-plus-search process supports PM retrieval in certain contexts, and that some discrepancy-plus-search mechanisms may result from the violation of processing expectations within a semantic context.

Age-Related Gene Expression Differences in Monocytes from Human Neonates, Young Adults, and Older Adults.

A variety of age-related differences in the innate and adaptive immune systems have been proposed to contribute to the increased susceptibility to infection of human neonates and older adults. The emergence of RNA sequencing (RNA-seq) provides an opportunity to obtain an unbiased, comprehensive, and quantitative view of gene expression differences in defined cell types from different age groups. An examination of ex vivo human monocyte responses to lipopolysaccharide stimulation or Listeria monocytogenes infection by RNA-seq revealed extensive similarities between neonates, young adults, and older adults, with an unexpectedly small number of genes exhibiting statistically significant age-dependent differences. By examining the differentially induced genes in the context of transcription factor binding motifs and RNA-seq data sets from mutant mouse strains, a previously described deficiency in interferon response factor-3 activity could be implicated in most of the differences between newborns and young adults. Contrary to these observations, older adults exhibited elevated expression of inflammatory genes at baseline, yet the responses following stimulation correlated more closely with those observed in younger adults. Notably, major differences in the expression of constitutively expressed genes were not observed, suggesting that the age-related differences are driven by environmental influences rather than cell-autonomous differences in monocyte development.

Infrared spectroscopy of Mg-CO2 and Al-CO2 complexes in helium nanodroplets.

The catalytic reduction of CO2 to produce hydrocarbon fuels is a topic that has gained significant attention. Development of efficient catalysts is a key enabler to such approaches, and metal-based catalysts have shown promise towards this goal. The development of a fundamental understanding of the interactions between CO2 molecules and metal atoms is expected to offer insight into the chemistry that occurs at the active site of such catalysts. In the current study, we utilize helium droplet methods to assemble complexes composed of a CO2 molecule and a Mg or Al atom. High-resolution infrared (IR) spectroscopy and optically selected mass spectrometry are used to probe the structure and binding of the complexes, and the experimental observations are compared with theoretical results determined from ab initio calculations. In both the Mg-CO2 and Al-CO2 systems, two IR bands are obtained: one assigned to a linear isomer and the other assigned to a T-shaped isomer. In the case of the Mg-CO2 complexes, the vibrational frequencies and rotational constants associated with the two isomers are in good agreement with theoretical values. In the case of the Al-CO2 complexes, the vibrational frequencies agree with theoretical predictions; however, the bands from both structural isomers exhibit significant homogeneous broadening sufficient to completely obscure the rotational structure of the bands. The broadening is consistent with an upper state lifetime of 2.7 ps for the linear isomer and 1.8 ps for the T-shaped isomer. The short lifetime is tentatively attributed to a prompt photo-induced chemical reaction between the CO2 molecule and the Al atom comprising the complex.

The selection and usage of information for perceiving and remembering intended and unintended object properties.

The current study was designed to investigate the linkage between intention, attention, and information selection and detection. Two experiments determined whether the intention to perceive maximum overhead reaching height with the use of handheld rods with different rotational inertia resulted in the ability to remember the rods' heaviness after they were removed from view. Participants were partially successful at distinguishing the heaviness of rods but only when visual information was restricted and reaching height was perceived using the rods by dynamic touch. The results support an ecological approach to attention, and suggest that information picked up for perception can support the memory of other properties of the environment if their informational bases are related.