DNA methylation-associated colonic mucosal immune and defense responses in treatment-naïve pediatric ulcerative colitis

Inflammatory bowel diseases (IBD) are emerging globally, indicating that environmental factors may be important in their pathogenesis. Colonic mucosal epigenetic changes, such as DNA methylation, can occur in response to the environment and have been implicated in IBD pathology. However, mucosal DNA methylation has not been examined in treatment-naïve patients. We studied DNA methylation in untreated, left sided colonic biopsy specimens using the Infinium HumanMethylation450 BeadChip array. We analyzed 22 control (C) patients, 15 untreated Crohn’s disease (CD) patients, and 9 untreated ulcerative colitis (UC) patients from two cohorts. Samples obtained at the time of clinical remission from two of the treatment-naïve UC patients were also included into the analysis. UC-specific gene expression was interrogated in a subset of adjacent samples (5 C and 5 UC) using the Affymetrix GeneChip PrimeView Human Gene Expression Arrays. Only treatment-naïve UC separated from control. One-hundred-and-twenty genes with significant expression change in UC (> 2-fold, P < 0.05) were associated with differentially methylated regions (DMRs). Epigenetically associated gene expression changes (including gene expression changes in the IFITM1, ITGB2, S100A9, SLPI, SAA1, and STAT3 genes) were linked to colonic mucosal immune and defense responses. These findings underscore the relationship between epigenetic changes and inflammation in pediatric treatment-naïve UC and may have potential etiologic, diagnostic, and therapeutic relevance for IBD

Nanomechanics combined with HDX reveals allosteric drug binding sites of CFTR NBD1.

Cystic fibrosis (CF) is a frequent genetic disease in Caucasians that is caused by the deletion of F508 (DF508) in the nucleotide binding domain 1 (NBD1) of the CF transmembrane conductance regulator (CFTR). The DF508 compromises the folding energetics of the NBD1, as well as the folding of three other CFTR domains. Combination of FDA approved corrector molecules can efficiently but incompletely rescue the DF508-CFTR folding and stability defect. Thus, new pharmacophores that would reinstate the wild-type-like conformational stability of the DF508-NBD1 would be highly beneficial. The most prominent molecule, 5-bromoindole-3-acetic acid (BIA) that can thermally stabilize the NBD1 has low potency and efficacy. To gain insights into the NBD1 (un)folding dynamics and BIA binding site localization, we combined molecular dynamics (MD) simulations, atomic force spectroscopy (AFM) and hydrogen- deuterium exchange (HDX) experiments. We found that the NBD1 a-subdomain with three adjacent strands from the b-subdomain plays an important role in early folding steps, when crucial non-native interactions are formed via residue F508. Our AFM and HDX experiments showed that BIA associates with this a-core region and increases the resistance of the DF508-NBD1 against mechanical unfolding, a phenomenon that could be exploited in future developments of folding correctors

Effective Area-Elasticity and Tension of Micro-manipulated Membranes

We evaluate the effective Hamiltonian governing, at the optically resolved scale, the elastic properties of micro-manipulated membranes. We identify floppy, entropic-tense and stretched-tense regimes, representing different behaviors of the effective area-elasticity of the membrane. The corresponding effective tension depends on the microscopic parameters (total area, bending rigidity) and on the optically visible area, which is controlled by the imposed external constraints. We successfully compare our predictions with recent data on micropipette experiments.Comment: To be published in Phys. Rev. Let

A Quantitative Theory of Mechanical Unfolding of a Homopolymer Globule

We propose the quantitative mean-field theory of mechanical unfolding of a globule formed by long flexible homopolymer chain collapsed in poor solvent and subjected to extensional deformation. We demonstrate that depending on the degree of polymerization and solvent quality (quantified by the Flory-Huggins $\chi$ parameter) the mechanical unfolding of the collapsed chain may either occur continuously (by passing a sequence of uniformly elongated configurations) or involves intra-molecular micro-phase coexistence of a collapsed and a stretched segment followed by an abrupt unraveling transition. The force-extension curves are obtained and quantitatively compared to our recent results of numerical self-consistent field (SCF) simulations. The phase diagrams for extended homopolymer chains in poor solvent comprising one- and two-phase regions are calculated for different chain length or/and solvent quality.Comment: 24 pages, 18 figure

Dynamics of folding in Semiflexible filaments

We investigate the dynamics of a single semiflexible filament, under the action of a compressing force, using numerical simulations and scaling arguments. The force is applied along the end to end vector at one extremity of the filament, while the other end is held fixed. We find that, unlike in elastic rods the filament folds asymmetrically with a folding length which depends only on the bending stiffness and the applied force. It is shown that this behavior can be attributed to the exponentially falling tension profile in the filament. While the folding time depends on the initial configuration, at late time, the distance moved by the terminal point of the filament and the length of the fold shows a power law dependence on time with an exponent 1/2.Comment: 13 pages, Late

Virus shapes and buckling transitions in spherical shells

We show that the icosahedral packings of protein capsomeres proposed by Caspar and Klug for spherical viruses become unstable to faceting for sufficiently large virus size, in analogy with the buckling instability of disclinations in two-dimensional crystals. Our model, based on the nonlinear physics of thin elastic shells, produces excellent one parameter fits in real space to the full three-dimensional shape of large spherical viruses. The faceted shape depends only on the dimensionless Foppl-von Karman number \gamma=YR^2/\kappa, where Y is the two-dimensional Young's modulus of the protein shell, \kappa is its bending rigidity and R is the mean virus radius. The shape can be parameterized more quantitatively in terms of a spherical harmonic expansion. We also investigate elastic shell theory for extremely large \gamma, 10^3 < \gamma < 10^8, and find results applicable to icosahedral shapes of large vesicles studied with freeze fracture and electron microscopy.Comment: 11 pages, 12 figure

Emergence of Collective Territorial Defense in Bacterial Communities: Horizontal Gene Transfer Can Stabilize Microbiomes

Multispecies bacterial communities such as the microbiota of the gastrointestinal tract can be remarkably stable and resilient even though they consist of cells and species that compete for resources and also produce a large number of antimicrobial agents. Computational modeling suggests that horizontal transfer of resistance genes may greatly contribute to the formation of stable and diverse communities capable of protecting themselves with a battery of antimicrobial agents while preserving a varied metabolic repertoire of the constituent species. In other words horizontal transfer of resistance genes makes a community compatible in terms of exoproducts and capable to maintain a varied and mature metagenome. The same property may allow microbiota to protect a host organism, or if used as a microbial therapy, to purge pathogens and restore a protective environment

Season of conception in rural gambia affects DNA methylation at putative human metastable epialleles.

Throughout most of the mammalian genome, genetically regulated developmental programming establishes diverse yet predictable epigenetic states across differentiated cells and tissues. At metastable epialleles (MEs), conversely, epigenotype is established stochastically in the early embryo then maintained in differentiated lineages, resulting in dramatic and systemic interindividual variation in epigenetic regulation. In the mouse, maternal nutrition affects this process, with permanent phenotypic consequences for the offspring. MEs have not previously been identified in humans. Here, using an innovative 2-tissue parallel epigenomic screen, we identified putative MEs in the human genome. In autopsy samples, we showed that DNA methylation at these loci is highly correlated across tissues representing all 3 embryonic germ layer lineages. Monozygotic twin pairs exhibited substantial discordance in DNA methylation at these loci, suggesting that their epigenetic state is established stochastically. We then tested for persistent epigenetic effects of periconceptional nutrition in rural Gambians, who experience dramatic seasonal fluctuations in nutritional status. DNA methylation at MEs was elevated in individuals conceived during the nutritionally challenged rainy season, providing the first evidence of a permanent, systemic effect of periconceptional environment on human epigenotype. At MEs, epigenetic regulation in internal organs and tissues varies among individuals and can be deduced from peripheral blood DNA. MEs should therefore facilitate an improved understanding of the role of interindividual epigenetic variation in human disease

Optical Trapping with High Forces Reveals Unexpected Behaviors of Prion Fibrils

Amyloid fibrils are important in diverse cellular functions, feature in many human diseases and have potential applications in nanotechnology. Here we describe methods that combine optical trapping and fluorescent imaging to characterize the forces that govern the integrity of amyloid fibrils formed by a yeast prion protein. A crucial advance was to use the self-templating properties of amyloidogenic proteins to tether prion fibrils, enabling their manipulation in the optical trap. At normal pulling forces the fibrils were impervious to disruption. At much higher forces (up to 250 pN), discontinuities occurred in force-extension traces before fibril rupture. Experiments with selective amyloid-disrupting agents and mutations demonstrated that such discontinuities were caused by the unfolding of individual subdomains. Thus, our results reveal unusually strong noncovalent intermolecular contacts that maintain fibril integrity even when individual monomers partially unfold and extend fibril length.National Institutes of Health (U.S.) (Grant GM025874)National Science Foundation (U.S.). CAREER (Award 0643745

Single Molecule Statistics and the Polynucleotide Unzipping Transition

We present an extensive theoretical investigation of the mechanical unzipping of double-stranded DNA under the influence of an applied force. In the limit of long polymers, there is a thermodynamic unzipping transition at a critical force value of order 10 pN, with different critical behavior for homopolymers and for random heteropolymers. We extend results on the disorder-averaged behavior of DNA's with random sequences to the more experimentally accessible problem of unzipping a single DNA molecule. As the applied force approaches the critical value, the double-stranded DNA unravels in a series of discrete, sequence-dependent steps that allow it to reach successively deeper energy minima. Plots of extension versus force thus take the striking form of a series of plateaus separated by sharp jumps. Similar qualitative features should reappear in micromanipulation experiments on proteins and on folded RNA molecules. Despite their unusual form, the extension versus force curves for single molecules still reveal remnants of the disorder-averaged critical behavior. Above the transition, the dynamics of the unzipping fork is related to that of a particle diffusing in a random force field; anomalous, disorder-dominated behavior is expected until the applied force exceeds the critical value for unzipping by roughly 5 pN.Comment: 40 pages, 18 figure