Synthesis, screening, and sequencing of cysteine-rich one-bead one-compound peptide libraries.

Cysteine-rich peptides are valued as tags for biarsenical fluorophores and as environmentally important reagents for binding toxic heavy metals. Due to the inherent difficulties created by cysteine, the power of one-bead one-compound (OBOC) libraries has never been applied to the discovery of short cysteine-rich peptides. We have developed the first method for the synthesis, screening, and sequencing of cysteine-rich OBOC peptide libraries. First, we synthesized a heavily biased cysteine-rich OBOC library, incorporating 50% cysteine at each position (Ac-X8-KM-TentaGel). Then, we developed conditions for cysteine alkylation, cyanogen bromide cleavage, and direct MS/MS sequencing of that library at the single bead level. The sequencing efficiency of this library was comparable to a traditional cysteine-free library. To validate screening of cysteine-rich OBOC libraries, we reacted a library with the biarsenical FlAsH and identified beads bearing the known biarsenical-binding motif (CCXXCC). These results enable OBOC libraries to be used in high-throughput discovery of cysteine-rich peptides for protein tagging, environmental remediation of metal contaminants, or cysteine-rich pharmaceuticals

The AST/RO Survey of the Galactic Center Region. I. The Inner 3 Degrees

We present fully-sampled maps of 461 GHz CO (4-3), 807 GHz CO (7-6), and 492 GHz [CI] (3P1-3P0) emission from the inner 3 degrees of the Galactic Center region taken with the Antarctic Submillimeter Telescope and Remote Observatory (AST/RO) in 2001--2002. The data cover -1.3 < l < 2, -0.3 < b < 0.2 with 0.5 arcmin spacing, resulting in spectra in 3 transitions at over 24,000 positions on the sky. The CO (4-3) emission is found to be essentially coextensive with lower-J transitions of CO. The CO (7-6) emission is spatially confined to a far smaller region than the lower-J CO lines. The [CI] (3P1-3P0) emission has a spatial extent similar to the low-J CO emission, but is more diffuse. Bright CO (7-6) emission is detected in the well-known Galactic Center clouds Sgr A and Sgr B. We also detect CO (4-3) and CO (7-6) absorption from spiral arms in the galactic disk at velocities near 0 km s^-1 along the line of sight to the Galactic Center. Analyzing our CO (7-6) and CO (4-3) data in conjunction with J = 1 - 0 12CO and 13CO data previously observed with the Bell Laboratories 7-m antenna, we apply a Large Velocity Gradient (LVG) model to estimate the kinetic temperature and density of molecular gas in the inner 200 pc of the Galactic Center region. We show maps of the derived distribution of gas density and kinetic temperature as a function of position and velocity for the entire region. Kinetic temperature was found to decrease from relatively high values (>70K) at cloud edges to low values (<50K) in the interiors. Typical gas pressures in the Galactic Center gas are n(H_2) T_kin approx 10^5.2 K cm^-3. We also present an (l,b) map of molecular hydrogen column density derived from our LVG results.Comment: 41 pages, 29 figures reduced to low resolution for astro-ph, for high resolution figures please go to http://cfa-www.harvard.edu/ASTRO/galcen_1.pdf this version incorporates changes due to accepted erratum (to appear in ApJS 153

Comparing personal insight gains due to consideration of a recent dream and consideration of a recent event using the Ullman and Schredl dream group methods

There have been reports and claims in the psychotherapeutic literature that the consideration of recent dreams can result in personal realizations and insight. There is theoretical support for these claims from work on rapid eye movement (REM) sleep having a function of the consolidation of emotional memories and the creative formation of connections between new and older memories. To investigate these claims, 11 participants (10 females, one male) reported and considered a recent home dream in a dream discussion group that following the “Appreciating dreams” method of Montague Ullman. The group ran 11 times, each participant attending and participating once. A further nine participants (seven females, two males) reported and considered a recent home dream in a group that followed the “Listening to the dreamer” method of Michael Schredl. The two studies each had a control condition where the participant also reported a recent event, the consideration of which followed the same technique as was followed for the dream report. Outcomes of the discussions were assessed by the participants on the Gains from Dream Interpretation (GDI) scale, and on its counterpart, the Gains from Event Interpretation scale. High ratings on the GDI experiential-insight subscale were reported for both methods, when applied to dreams, and for the Ullman method Exploration-Insight ratings for the dream condition were significantly higher than for the control event condition. In the Ullman method, self-assessment of personal insight due to consideration of dream content was also significantly higher than for the event consideration condition. The findings support the view that benefits can be obtained from the consideration of dream content, in terms of identifying the waking life sources of dream content, and because personal insight may also occur. To investigate the mechanisms for the findings, the studies should be repeated with REM and non-REM dream reports, hypothesizing greater insight from the former

Optical Colors of Intracluster Light in the Virgo Cluster Core

We continue our deep optical imaging survey of the Virgo cluster using the CWRU Burrell Schmidt telescope by presenting B-band surface photometry of the core of the Virgo cluster in order to study the cluster's intracluster light (ICL). We find ICL features down to mu_b ~ 29 mag sq. arcsec, confirming the results of Mihos et al. (2005), who saw a vast web of low-surface brightness streams, arcs, plumes, and diffuse light in the Virgo cluster core using V-band imaging. By combining these two data sets, we are able to measure the optical colors of many of the cluster's low-surface brightness features. While much of our imaging area is contaminated by galactic cirrus, the cluster core near the cD galaxy, M87, is unobscured. We trace the color profile of M87 out to over 2000 arcsec, and find a blueing trend with radius, continuing out to the largest radii. Moreover, we have measured the colors of several ICL features which extend beyond M87's outermost reaches and find that they have similar colors to the M87's halo itself, B-V ~ 0.8. The common colors of these features suggests that the extended outer envelopes of cD galaxies, such as M87, may be formed from similar streams, created by tidal interactions within the cluster, that have since dissolved into a smooth background in the cluster potential.Comment: 14 pages. Published in ApJ, September 201

Southern Hemisphere controls on ITCZ variability in southwest Madagascar over the past 117,000 years

Migration of the inter-tropical convergence zone, driven by changes in seasonal insolation and high northern latitude temperatures, is the primary control on tropical rainfall on geologic timescales. We test this paradigm using the timing of growth of stalagmites from southwest Madagascar to infer the timing of expansion of the ITCZ to the south at its southern limit. Over the past 117 ky, speleothems grew in the study area primarily when two conditions are met: summer insolation greater than the mean and relatively high Southern Hemisphere temperatures as indicted by maxima in Antarctic ice core oxygen isotope ratios. We observe little influence of Northern Hemisphere, millennial scale temperature variability on the pluvial periods. Further, we observe periods during which the ITCZ simultaneously expands or contracts in both hemispheres. Because Antarctic isotope maxima are periods of increased atmospheric CO2, our results have implications for how tropical rainfall in the Southern Hemisphere might respond to global warming

Loss of histone methyltransferase Ezh2 stimulates an osteogenic transcriptional program in chondrocytes but does not affectcartilage development

Ezh2 is a histone methyltransferase that suppresses osteoblast maturation and skeletal development. We evaluated the roleof Ezh2 in chondrocyte lineage differentiation and endochondral ossification. Ezh2 was genetically inactivated in the mesenchymal, osteoblastic, and chondrocytic lineages in mice using the Prrx1-Cre,Osx1-Cre, and Col2a1-Cre drivers, respectively. Wild-type and conditional knockout mice were phenotypically assessed by grossmorphology, histology, and micro-CT imaging. Ezh2-deficient chondrocytes in micromass culture models were evaluated usingRNA-sequencing, histologic evaluation, and western blotting. Aged mice with Ezh2 deficiency were also evaluated for prematuredevelopment of osteoarthritis using radiographic analysis. Ezh2 deficiency in murine chondrocytes reduced bone density at 4 weeks of age, although caused no other gross developmentaleffects. Knockdown of Ezh2 in chondrocyte micromass cultures resulted in a global reduction in trimethylation of histone 3lysine 27 (H3K27me3) and altered differentiation in vitro. RNA-seq analysis revealed enrichment of an osteogenic gene expressionprofile in Ezh2 deficient chondrocytes. Joint development proceeded normally in the absence of Ezh2 in chondrocytes withoutinducing excessive hypertrophy or premature osteoarthritis in vivo. In summary, loss of Ezh2 reduced H3K27me3 levels, increased expression of osteogenic genes in chondrocytes, and resulted ina transient post-natal bone phenotype. Remarkably, Ezh2 activity is dispensable for normal chondrocyte maturation and endochondralossification in vivo, even though it appears to have a critical role during early stages of mesenchymal lineage-commitment

Differences in genotype and virulence among four multidrug-resistant <i>Streptococcus pneumoniae</i> isolates belonging to the PMEN1 clone

We report on the comparative genomics and characterization of the virulence phenotypes of four &lt;i&gt;S. pneumoniae&lt;/i&gt; strains that belong to the multidrug resistant clone PMEN1 (Spain&lt;sup&gt;23F&lt;/sup&gt; ST81). Strains SV35-T23 and SV36-T3 were recovered in 1996 from the nasopharynx of patients at an AIDS hospice in New York. Strain SV36-T3 expressed capsule type 3 which is unusual for this clone and represents the product of an in vivo capsular switch event. A third PMEN1 isolate - PN4595-T23 - was recovered in 1996 from the nasopharynx of a child attending day care in Portugal, and a fourth strain - ATCC700669 - was originally isolated from a patient with pneumococcal disease in Spain in 1984. We compared the genomes among four PMEN1 strains and 47 previously sequenced pneumococcal isolates for gene possession differences and allelic variations within core genes. In contrast to the 47 strains - representing a variety of clonal types - the four PMEN1 strains grouped closely together, demonstrating high genomic conservation within this lineage relative to the rest of the species. In the four PMEN1 strains allelic and gene possession differences were clustered into 18 genomic regions including the capsule, the blp bacteriocins, erythromycin resistance, the MM1-2008 prophage and multiple cell wall anchored proteins. In spite of their genomic similarity, the high resolution chinchilla model was able to detect variations in virulence properties of the PMEN1 strains highlighting how small genic or allelic variation can lead to significant changes in pathogenicity and making this set of strains ideal for the identification of novel virulence determinant

An Atlas of FUSE Sight Lines Toward the Magellanic Clouds

We present an atlas of 57 Large Magellanic Cloud (LMC) and 37 Small Magellanic Cloud (SMC) observations obtained with the Far Ultraviolet Spectroscopic Explorer (FUSE) satellite. The atlas highlights twelve interstellar absorption line transitions at a resolution of ~15 km/s. These transitions cover a broad range of temperatures, ionization states, and abundances. The species included are OVI, which probes hot (T~3x10^5 K) ionized gas; CIII and FeIII, which probe warm (T~10^4 K) ionized gas; SiII, PII, CII, FeII, and OI, warm neutral gas; and six different molecular hydrogen transitions, which trace cold (T<=500 K) gas. We include Schmidt Halpha CCD images of the region surrounding each sight line showing the morphology of warm ionized gas in the vicinity, along with continuum images near each FUSE aperture position. Finally, we present several initial scientific results derived from this dataset on the interstellar medium of the Magellanic Clouds and Galactic halo.Comment: 29 pages, 6 figures. Complete Atlas of 94 additional images (~800kB each) is available at http://fuse.pha.jhu.edu/~danforth/atlas Accepted to the ApJS March 200

Structure of an Enzyme-Derived Phosphoprotein Recognition Domain

Membrane Associated Guanylate Kinases (MAGUKs) contain a protein interaction domain (GKdom) derived from the enzyme Guanylate Kinase (GKenz). Here we show that GKdom from the MAGUK Discs large (Dlg) is a phosphoprotein recognition domain, specifically recognizing the phosphorylated form of the mitotic spindle orientation protein Partner of Inscuteable (Pins). We determined the structure of the Dlg-Pins complex to understand the dramatic transition from nucleotide kinase to phosphoprotein recognition domain. The structure reveals that the region of the GKdom that once served as the GMP binding domain (GBD) has been co-opted for protein interaction. Pins makes significantly more contact with the GBD than does GMP, but primarily with residues that are conserved between enzyme and domain revealing the versatility of the GBD as a platform for nucleotide and protein interactions. Mutational analysis reveals that the GBD is also used to bind the GK ligand MAP1a, suggesting that this is a common mode of MAGUK complex assembly. The GKenz undergoes a dramatic closing reaction upon GMP binding but the protein-bound GKdom remains in the ‘open’ conformation indicating that the dramatic conformational change has been lost in the conversion from nucleotide kinase to phosphoprotein recognition domain

Prediction of enteric methane production, yield and intensity in dairy cattle using an intercontinental database

Enteric methane (CH4) production from cattle contributes to global greenhouse gas emissions. Measurement of enteric CH4 is complex, expensive and impractical at large scales; therefore, models are commonly used to predict CH4 production. However, building robust prediction models requires extensive data from animals under different management systems worldwide. The objectives of this study were to (1) collate a global database of enteric CH4 production from individual lactating dairy cattle; (2) determine the availability of key variables for predicting enteric CH4 production (g/d per cow), yield [g/kg dry matter intake (DMI)], and intensity (g/kg energy corrected milk) and their respective relationships; (3) develop intercontinental and regional models and cross-validate their performance; and (4) assess the trade-off between availability of on-farm inputs and CH4 prediction accuracy. The intercontinental database covered Europe (EU), the US (US), Chile (CL), Australia (AU), and New Zealand (NZ). A sequential approach was taken by incrementally adding key variables to develop models with increasing complexity. Methane emissions were predicted by fitting linear mixed models. Within model categories, an intercontinental model with the most available independent variables performed best with root mean square prediction error (RMSPE) as a percentage of mean observed value of 16.6, 14.4, and 19.8% for intercontinental, EU, and US regions, respectively. Less complex models requiring only DMI had predictive ability comparable to complex models. Enteric CH4 production, yield, and intensity prediction models developed on an intercontinental basis had similar performance across regions, however, intercepts and slopes were different with implications for prediction. Revised CH4 emission conversion factors for specific regions are required to improve CH4 production estimates in national inventories. In conclusion, information on DMI is required for good prediction, and other factors such as dietary NDF concentration, improve the prediction. For enteric CH4 yield and intensity prediction, information on milk yield and composition is required for better estimation