Targeting the Ets Binding Site of the HER2/neu Promoter with Pyrrole-Imidazole Polyamides

Three DNA binding polyamides (1-3) were synthesized that bind with high affinity (Ka = 8.7·10^9 M^-1 to 1.4·10^10 M^-1) to two 7-base pair sequences overlapping the Ets DNA binding site (EBS; GAGGAA) within the regulatory region of the HER2/neu proximal promoter. As measured by electrophoretic mobility shift assay, polyamides binding to flanking elements upstream (1) or downstream (2 and 3) of the EBS were one to two orders of magnitude more effective than the natural product distamycin at inhibiting formation of complexes between the purified EBS protein, epithelial restricted with serine box (ESX), and the HER2/neu promoter probe. One polyamide, 2, completely blocked Ets-DNA complex formation at 10 nM ligand concentration, whereas formation of activator protein-2-DNA complexes was unaffected at the activator protein-2 binding site immediately upstream of the HER2/neu EBS, even at 100 nM ligand concentration. At equilibrium, polyamide 1 was equally effective at inhibiting Ets/DNA binding when added before or after in vitro formation of protein-promoter complexes, demonstrating its utility to disrupt endogenous Ets-mediated HER2/neu preinitiation complexes. Polyamide 2, the most potent inhibitor of Ets-DNA complex formation by electrophoretic mobility shift assay, was also the most effective inhibitor of HER2/neu promoter-driven transcription measured in a cell-free system using nuclear extract from an ESX- and HER2/neu-overexpressing human breast cancer cell line, SKBR-3

A genome-wide linkage scan for body mass index on Framingham Heart Study families

BACKGROUND: Genome-wide scan data from a community-based sample was used to identify the genetic factors that affect body mass index (BMI). BMI was defined as weight (kg) over the square of height (m), where weight and height were obtained from the first measurement available between the ages of 40 and 50 years. RESULTS: Significant familial correlations were observed in mother:father (spouse) relative pairs and in all relative pairs examined except parent:daughter pairs. Single-point sib-pair regression analysis provided nominal evidence for linkage (p < 0.05) of loci to BMI at 23 markers. Multi-point sib-pair regression analysis provided nominal evidence for linkage to BMI at 42 loci on 12 chromosomes. Empirical p-values showed results consistent with the multi-point results; all but three of the loci identified by multi-point analysis were also significant. CONCLUSION: The largest regions of nominally significant linkage were found on chromosomes 2, 3, and 11. The most significant evidence for linkage was obtained with markers D2S1788, D2S1356, D2S1352, D3S1744, and D11S912 from multi-point sib-pair single-trait regression analysis. Our results are in agreement with some of the recently published reports on BMI using various data sets including the Framingham Heart Study data

Litigation: Time to Revisit \u3cem\u3eChevron\u3c/em\u3e Deference?

This panel discussion took place on Thursday, November 13, 2014 at the Mayflower Hotel in Washington, D.C., prior to the passing of Justice Antonin Scalia. Justice Scalia\u27s impact on the development of administrative law in the United States is unparalleled

Genomic regions linked to alcohol consumption in the Framingham Heart Study

BACKGROUND: Pedigree, demographic, square-root transformed maximum alcohol (SRMAXAPD) and maximum cigarette (MAXCPD) consumption, and genome-wide scan data from the Framingham Heart Study (FHS) were used to investigate genetic factors that may affect alcohol and cigarette consumption in this population-based sample. RESULTS: A significant sister:sister correlation greater than spouse correlation was observed for MAXCPD only. Single-point sib-pair regression analysis provided nominal evidence for linkage of loci to both SRMAXAPD and MAXCPD consumption traits, with more significant evidence of linkage to SRMAXAPD than to MAXCPD. One genomic region, chr9q21.11, exhibits significant multi-point sib-pair regression to SRMAXAPD. CONCLUSION: SRMAXAPD exhibits greater evidence for genetic linkage than does MAXCPD in the FHS sample. Four regions of the genome exhibiting nominal evidence for linkage to SRMAXAPD in the FHS sample correspond to regions of the genome previously identified as linked to alcoholism or related traits in the family data set ascertained on individuals affected with alcohol dependence known as COGA

Performance of high-throughput DNA quantification methods

BACKGROUND: The accuracy and precision of estimates of DNA concentration are critical factors for efficient use of DNA samples in high-throughput genotype and sequence analyses. We evaluated the performance of spectrophotometric (OD) DNA quantification, and compared it to two fluorometric quantification methods, the PicoGreen(® )assay (PG), and a novel real-time quantitative genomic PCR assay (QG) specific to a region at the human BRCA1 locus. Twenty-Two lymphoblastoid cell line DNA samples with an initial concentration of ~350 ng/uL were diluted to 20 ng/uL. DNA concentration was estimated by OD and further diluted to 5 ng/uL. The concentrations of multiple aliquots of the final dilution were measured by the OD, QG and PG methods. The effects of manual and robotic laboratory sample handling procedures on the estimates of DNA concentration were assessed using variance components analyses. RESULTS: The OD method was the DNA quantification method most concordant with the reference sample among the three methods evaluated. A large fraction of the total variance for all three methods (36.0–95.7%) was explained by sample-to-sample variation, whereas the amount of variance attributable to sample handling was small (0.8–17.5%). Residual error (3.2–59.4%), corresponding to un-modelled factors, contributed a greater extent to the total variation than the sample handling procedures. CONCLUSION: The application of a specific DNA quantification method to a particular molecular genetic laboratory protocol must take into account the accuracy and precision of the specific method, as well as the requirements of the experimental workflow with respect to sample volumes and throughput. While OD was the most concordant and precise DNA quantification method in this study, the information provided by the quantitative PCR assay regarding the suitability of DNA samples for PCR may be an essential factor for some protocols, despite the decreased concordance and precision of this method

Revealing the Structure of a Pre-Transitional Disk: The Case of the Herbig F Star SAO 206462 (HD 135344B)

SAO 206462 (HD 135344B) has previously been identified as a Herbig F star with a circumstellar disk with a dip in its infrared excess near 10 mum. In combination with a low accretion rate estimated from Br gamma, it may represent a gapped, but otherwise primordial or pre-transitional disk. We test this hypothesis with Hubble Space Telescope coronagraphic imagery, FUV spectroscopy and imagery and archival X-ray data, and spectral energy distribution (SED) modeling constrained by the observed system inclination, disk outer radius, and outer disk radial surface brightness (SB) profile using the Whitney Monte Carlo Radiative Transfer Code. The essentially face-on (i lsim 20°) disk is detected in scattered light from 0farcs4 to 1farcs15 (56-160 AU), with a steep (r -9.6) radial SB profile from 0farcs6 to 0farcs93. Fitting the SB data requires a concave upward or anti-flared outer disk, indicating substantial dust grain growth and settling by 8 ± 4 Myr. The warm dust component is significantly variable in near to mid-IR excess and in temperature. At its warmest, it appears confined to a narrow belt from 0.08 to 0.2 AU. The steep SED for this dust component is consistent with grains with a\u3c= 2.5 mum. For cosmic carbon to silicate dust composition, conspicuous 10 mum silicate emission would be expected and is not observed. This may indicate an elevated carbon to silicate ratio for the warm dust, which is not required to fit the outer disk. At its coolest, the warm dust can be fit with a disk from 0.14 to 0.31 AU, but with a higher inclination than either the outer disk or the gaseous disk, providing confirmation of the high inclination inferred from mid-IR interferometry. In tandem, the compositional and inclination difference between the warm dust and the outer dust disk suggests that the warm dust may be of second-generation origin, rather than a remnant of a primordial disk component. With its near face-on inclination, SAO 206462\u27s disk is a prime location for planet searches

Relating jet structure to photometric variability: the Herbig Ae star HD 163296

Herbig Ae/Be stars are intermediate-mass pre-main sequence stars surrounded by circumstellar dust disks. Some are observed to produce jets, whose appearance as a sequence of shock fronts (knots) suggests a past episodic outflow variability. This "jet fossil record" can be used to reconstruct the outflow history. We present the first optical to near-infrared (NIR) VLT/X-shooter spectra of the jet from the Herbig Ae star HD 163296. We determine physical conditions in the knots, as well as their kinematic "launch epochs". Knots are formed simultaneously on either side of the disk, with a regular interval of ~16 yr. The velocity dispersion versus jet velocity and the energy input are comparable in both lobes. However, the mass loss rate, velocity, and shock conditions are asymmetric. We find Mjet/Macc ~ 0.01-0.1, consistent with magneto-centrifugal jet launching models. No evidence for dust is found in the high-velocity jet, suggesting it is launched within the sublimation radius (<0.5 au). The jet inclination measured from proper motions and radial velocities confirms it is perpendicular to the disk. A tentative relation is found between the structure of the jet and the photometric variability of the source. Episodes of NIR brightening were previously detected and attributed to a dusty disk wind. We report for the first time significant optical fadings lasting from a few days up to a year, coinciding with the NIR brightenings. These are likely caused by dust lifted high above the disk plane; this supports the disk wind scenario. The disk wind is launched at a larger radius than the high-velocity atomic jet, although their outflow variability may have a common origin. No significant relation between outflow and accretion variability could be established. Our findings confirm that this source undergoes periodic ejection events, which may be coupled with dust ejections above the disk plane.Comment: 20 pages, 11 figures, accepted for publication in Astronomy & Astrophysic