During B lymphocyte development, immunoglobulin heavy-chain variable (VH), diversity (DH), and joining (JH) segments assemble to generate a diverse antigen receptor repertoire. Here, we have marked ...the distal VH and DH-JH-Eμ regions with Tet-operator binding sites and traced their 3D trajectories in pro-B cells transduced with a retrovirus encoding Tet-repressor-EGFP. We found that these elements displayed fractional Langevin motion (fLm) due to the viscoelastic hindrance from the surrounding network of proteins and chromatin fibers. Using fractional Langevin dynamics modeling, we found that, with high probability, DHJH elements reach a VH element within minutes. Spatial confinement emerged as the dominant parameter that determined the frequency of such encounters. We propose that the viscoelastic nature of the nuclear environment causes coding elements and regulatory elements to bounce back and forth in a spring-like fashion until specific genomic interactions are established and that spatial confinement of topological domains largely controls first-passage times for genomic interactions.
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•3D trajectories adopted by the immunoglobulin heavy-chain locus•Fractional Langevin motion and VH-DHJH genomic interactions•First-passage times for coding and regulatory DNA elements•Spatial confinement largely controls encounter times for genomic interactions
Spatial confinement, related to the viscoelastic nature of the nuclear environment, emerges as the dominant factor that determines how frequent and how fast coding and regulatory sequences of the genome interact with each other.
Perfluorooctane sulfonic acid (PFOS) is a long chain per- and polyfluoroalklyl substance (PFAS) that has been used in aqueous film-forming foams. Emerging epidemiological evidence indicates that PFOS ...may be associated with chronic lung diseases such as asthma and analysis of human tissues demonstrates that the lungs carry a significant body burden of PFOS. Deficits in barrier function are a major risk factor for asthma. Thus, we hypothesized that PFOS exposure will lead to impaired epithelial barrier function through dysregulated tight junctions. Hence, we assessed the impact of PFOS on epithelial barrier integrity. Bronchial epithelial cells (16HBE) were grown on collagen-coated transwells and treated to 5-25 μM PFOS, and assessed for changes in barrier function and tight junction proteins. Rescue experiments were performed using the protein kinase D (PKD) inhibitor, CID755673. PFOS treatment reduced transepithelial electrical resistance (TEER) and increased 4 kDa FITC-dextran flux. Additionally, PFOS significantly decreased protein levels and the tight junction organization rate of occludin and zonula occludens 1. Increased phosphorylation (Ser744/Ser748) of PKD was observed 3 h following PFOS treatment. Pretreatment with the PKD inhibitor attenuated PFOS-mediated changes in TEER and FITC-dextran flux and restored occludin protein levels. In conclusion, PFOS causes loss of airway barrier integrity and the disruption of tight junctions in bronchial epithelial cells, which was partly attenuated through the inhibition of PKD. These findings demonstrate that PFOS is capable of disrupting airway barrier function, a potentially driving factor underlying associations between PFOS and respiratory diseases such as asthma.
Diverse antibody repertoires are generated through remote genomic interactions involving immunoglobulin variable (V
), diversity (D
) and joining (J
) gene segments. How such interactions are ...orchestrated remains unknown. Here we develop a strategy to track V
-D
J
motion in B-lymphocytes. We find that V
and D
J
segments are trapped in configurations that allow only local motion, such that spatially proximal segments remain in proximity, while spatially remote segments remain remote. Within a subset of cells, however, abrupt changes in V
-D
J
motion are observed, plausibly caused by temporal alterations in chromatin configurations. Comparison of experimental and simulated data suggests that constrained motion is imposed by a network of cross-linked chromatin chains characteristic of a gel phase, yet poised near the sol phase, a solution of independent chromatin chains. These results suggest that chromosome organization near the sol-gel phase transition dictates the timing of genomic interactions to orchestrate gene expression and somatic recombination.
There is great potential for host-based gene expression analysis to impact the early diagnosis of infectious diseases. In particular, the influenza pandemic of 2009 highlighted the challenges and ...limitations of traditional pathogen-based testing for suspected upper respiratory viral infection. We inoculated human volunteers with either influenza A (A/Brisbane/59/2007 (H1N1) or A/Wisconsin/67/2005 (H3N2)), and assayed the peripheral blood transcriptome every 8 hours for 7 days. Of 41 inoculated volunteers, 18 (44%) developed symptomatic infection. Using unbiased sparse latent factor regression analysis, we generated a gene signature (or factor) for symptomatic influenza capable of detecting 94% of infected cases. This gene signature is detectable as early as 29 hours post-exposure and achieves maximal accuracy on average 43 hours (p = 0.003, H1N1) and 38 hours (p-value = 0.005, H3N2) before peak clinical symptoms. In order to test the relevance of these findings in naturally acquired disease, a composite influenza A signature built from these challenge studies was applied to Emergency Department patients where it discriminates between swine-origin influenza A/H1N1 (2009) infected and non-infected individuals with 92% accuracy. The host genomic response to Influenza infection is robust and may provide the means for detection before typical clinical symptoms are apparent.
The tumor microenvironment has a significant impact on tumor development. Two important determinants in this environment are hypoxia and lactic acidosis. Although lactic acidosis has long been ...recognized as an important factor in cancer, relatively little is known about how cells respond to lactic acidosis and how that response relates to cancer phenotypes. We develop genome-scale gene expression studies to dissect transcriptional responses of primary human mammary epithelial cells to lactic acidosis and hypoxia in vitro and to explore how they are linked to clinical tumor phenotypes in vivo. The resulting experimental signatures of responses to lactic acidosis and hypoxia are evaluated in a heterogeneous set of breast cancer datasets. A strong lactic acidosis response signature identifies a subgroup of low-risk breast cancer patients having distinct metabolic profiles suggestive of a preference for aerobic respiration. The association of lactic acidosis response with good survival outcomes may relate to the role of lactic acidosis in directing energy generation toward aerobic respiration and utilization of other energy sources via inhibition of glycolysis. This "inhibition of glycolysis" phenotype in tumors is likely caused by the repression of glycolysis gene expression and Akt inhibition. Our study presents a genomic evaluation of the prognostic information of a lactic acidosis response independent of the hypoxic response. Our results identify causal roles of lactic acidosis in metabolic reprogramming, and the direct functional consequence of lactic acidosis pathway activity on cellular responses and tumor development. The study also demonstrates the utility of genomic analysis that maps expression-based findings from in vitro experiments to human samples to assess links to in vivo clinical phenotypes.
Perfluorooctane sulfonic acid (PFOS) is a long-chain per- and polyfluoroalkyl substance (PFAS), a persistent organic pollutant, which has been used in aqueous film-forming foams. Emerging ...epidemiological evidence indicates a significant body burden of PFOS is observed in the lungs. Furthermore, developmental PFOS exposure dysregulates lung development and exacerbates eosinophilic inflammation, which are critical risk factors for asthma. However, it is unknown whether PFOS exerts sex-dependent effects on house dust mite (HDM) induced asthmatic progression and allergic inflammation. In this study, timed pregnant Balb/cJ dams were dosed orally via PFOS (1.0 mg/kg/d) spiked or vehicle control mealworms from gestational day (GD) 0.5 to postnatal day (PND) 21. Subsequently, HDM (30 μg/day) was administered starting at PND 77–82 for 10 days, and the mice were sacrificed 48 h after their final treatment. The serum and lung PFOS concentrations were 3.391 ± 0.189 μg/mL and 3.567 ± 0.1676 μg/g in the offspring, respectively. Male mice exposed to PFOS + HDM showed higher total cell counts in bronchoalveolar lavage fluid (BALF), macrophage counts, and eosinophil counts compared to mice exposed to HDM alone. Female mice exposed to PFOS + HDM had increased BALF eosinophil percentage, mucous production, alternatively activated (M2) macrophage polarization, and M2-associated gene expression compared to female mice exposed to HDM alone. PFOS exposure had no significant effect on HDM-induced IL-4, IL-5, or IL-13, but RANTES was further elevated in female mice. Overall, our data suggest that developmental PFOS exposure increased the risk of exacerbated eosinophilic inflammation and M2 polarization, which were more severe in female mice, suggesting sex-dependent developmental effects of PFOS on allergic airway responses.
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•Perfluorooctane sulfonic acid (PFOS) accumulated in the lungs of offspring following developmental exposure.•Developmental PFOS exposure exhibited sex-dependent effects on house dust mite (HDM) induced allergic inflammation and asthmatic progression in adult offspring.•PFOS exacerbated eosinophilic inflammation and M2 polarization, which were more severe in female mice.•PFOS exposure potentiated type 1 interferon signaling in HDM exposed male mice.
We describe studies in molecular profiling and biological pathway analysis that use sparse latent factor and regression models for microarray gene expression data. We discuss breast cancer ...applications and key aspects of the modeling and computational methodology. Our case studies aim to investigate and characterize heterogeneity of structure related to specific oncogenic pathways, as well as links between aggregate patterns in gene expression profiles and clinical biomarkers. Based on the metaphor of statistically derived "factors" as representing biological "subpathway" structure, we explore the decomposition of fitted sparse factor models into pathway subcomponents and investigate how these components overlay multiple aspects of known biological activity. Our methodology is based on sparsity modeling of multivariate regression, ANOVA, and latent factor models, as well as a class of models that combines all components. Hierarchical sparsity priors address questions of dimension reduction and multiple comparisons, as well as scalability of the methodology. The models include practically relevant non-Gaussian/nonparametric components for latent structure, underlying often quite complex non-Gaussianity in multivariate expression patterns. Model search and fitting are addressed through stochastic simulation and evolutionary stochastic search methods that are exemplified in the oncogenic pathway studies. Supplementary supporting material provides more details of the applications, as well as examples of the use of freely available software tools for implementing the methodology.
A conserved segment, i.e. a segment of chromosome unbroken during evolution, is an important operational concept in comparative genomics. Until now, algorithms that are designed to identify conserved ...segments often return synteny blocks that overlap, synteny blocks that include micro-rearrangements or synteny blocks erroneously short. Here we present definitions of conserved segments and synteny blocks independent of any heuristic method and we describe four new post-processing strategies to refine synteny blocks into accurate conserved segments. The first strategy identifies micro-rearrangements, the second strategy identifies mono-genic conserved segments, the third returns non-overlapping segments and the fourth repairs incorrect ruptures of synteny. All these refinements are implemented in a new version of PhylDiag that has been benchmarked against i-ADHoRe 3.0 and Cyntenator, based on a realistic simulated evolution and true simulated conserved segments.
Little is known about the reproductive effects of paternal cannabis exposure. We evaluated associations between cannabis or tetrahydrocannabinol (THC) exposure and altered DNA methylation in sperm ...from humans and rats, respectively. DNA methylation, measured by reduced representation bisulfite sequencing, differed in the sperm of human users from non-users by at least 10% at 3,979 CpG sites. Pathway analyses indicated Hippo Signaling and Pathways in Cancer as enriched with altered genes (Bonferroni p < 0.02). These same two pathways were also enriched with genes having altered methylation in sperm from THC-exposed versus vehicle-exposed rats (p < 0.01). Data validity is supported by significant correlations between THC exposure levels in humans and methylation for 177 genes, and substantial overlap in THC target genes in rat sperm (this study) and genes previously reported as having altered methylation in the brain of rat offspring born to parents both exposed to THC during adolescence. In humans, cannabis use was also associated with significantly lower sperm concentration. Findings point to possible pre-conception paternal reproductive risks associated with cannabis use.
Sex differences in Alzheimer's disease (AD) biology and progression are not yet fully characterized. The goal of this study is to examine the effect of sex on cognitive progression in subjects with ...high likelihood of mild cognitive impairment (MCI) due to Alzheimer's and followed up to 10 years in the Alzheimer's Disease Neuroimaging Initiative (ADNI). Cerebrospinal fluid total-tau and amyloid-beta (Aβ42) ratio values were used to sub-classify 559 MCI subjects (216 females, 343 males) as having "high" or "low" likelihood for MCI due to Alzheimer's. Data were analyzed using mixed-effects models incorporating all follow-ups. The worsening from baseline in Alzheimer's Disease Assessment Scale-Cognitive score (mean, SD) (9 ± 12) in subjects with high likelihood of MCI due to Alzheimer's was markedly greater than that in subjects with low likelihood (1 ± 6, p < 0.0001). Among MCI due to AD subjects, the mean worsening in cognitive score was significantly greater in females (11.58 ± 14) than in males (6.87 ± 11, p = 0.006). Our findings highlight the need to further investigate these findings in other populations and develop sex specific timelines for Alzheimer's disease progression.
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