Genome-Wide Modeling of Transcription Preinitiation Complex Disassembly Mechanisms using ChIP-chip Data

Apparent occupancy levels of proteins bound to DNA in vivo can now be routinely measured on a genomic scale. A challenge in relating these occupancy levels to assembly mechanisms that are defined with biochemically isolated components lies in the veracity of assumptions made regarding the in vivo system. Assumptions regarding behavior of molecules in vivo can neither be proven true nor false, and thus is necessarily subjective. Nevertheless, within those confines, connecting in vivo protein-DNA interaction observations with defined biochemical mechanisms is an important step towards fully defining and understanding assembly/disassembly mechanisms in vivo. To this end, we have developed a computational program PathCom that models in vivo protein-DNA occupancy data as biochemical mechanisms under the assumption that occupancy levels can be related to binding duration and explicitly defined assembly/disassembly reactions. We exemplify the process with the assembly of the general transcription factors (TBP, TFIIB, TFIIE, TFIIF, TFIIH, and RNA polymerase II) at the genes of the budding yeast Saccharomyces. Within the assumption inherent in the system our modeling suggests that TBP occupancy at promoters is rather transient compared to other general factors, despite the importance of TBP in nucleating assembly of the preinitiation complex. PathCom is suitable for modeling any assembly/disassembly pathway, given that all the proteins (or species) come together to form a complex

Update on the correlation of the highest energy cosmic rays with nearby extragalactic matter

Data collected by the Pierre Auger Observatory through 31 August 2007 showed evidence for anisotropy in the arrival directions of cosmic rays above the Greisen-Zatsepin-Kuz'min energy threshold, \nobreak{$6\times 10^{19}$eV}. The anisotropy was measured by the fraction of arrival directions that are less than $3.1^\circ$ from the position of an active galactic nucleus within 75 Mpc (using the V\'eron-Cetty and V\'eron $12^{\rm th}$ catalog). An updated measurement of this fraction is reported here using the arrival directions of cosmic rays recorded above the same energy threshold through 31 December 2009. The number of arrival directions has increased from 27 to 69, allowing a more precise measurement. The correlating fraction is $(38^{+7}_{-6})$, compared with $21$ expected for isotropic cosmic rays. This is down from the early estimate of $(69^{+11}_{-13})$. The enlarged set of arrival directions is examined also in relation to other populations of nearby extragalactic objects: galaxies in the 2 Microns All Sky Survey and active galactic nuclei detected in hard X-rays by the Swift Burst Alert Telescope. A celestial region around the position of the radiogalaxy Cen A has the largest excess of arrival directions relative to isotropic expectations. The 2-point autocorrelation function is shown for the enlarged set of arrival directions and compared to the isotropic expectation.Comment: Accepted for publication in Astroparticle Physics on 31 August 201

The Fluorescence Detector of the Pierre Auger Observatory

The Pierre Auger Observatory is a hybrid detector for ultra-high energy cosmic rays. It combines a surface array to measure secondary particles at ground level together with a fluorescence detector to measure the development of air showers in the atmosphere above the array. The fluorescence detector comprises 24 large telescopes specialized for measuring the nitrogen fluorescence caused by charged particles of cosmic ray air showers. In this paper we describe the components of the fluorescence detector including its optical system, the design of the camera, the electronics, and the systems for relative and absolute calibration. We also discuss the operation and the monitoring of the detector. Finally, we evaluate the detector performance and precision of shower reconstructions.Comment: 53 pages. Submitted to Nuclear Instruments and Methods in Physics Research Section

Advanced functionality for radio analysis in the Offline software framework of the Pierre Auger Observatory

The advent of the Auger Engineering Radio Array (AERA) necessitates the development of a powerful framework for the analysis of radio measurements of cosmic ray air showers. As AERA performs "radio-hybrid" measurements of air shower radio emission in coincidence with the surface particle detectors and fluorescence telescopes of the Pierre Auger Observatory, the radio analysis functionality had to be incorporated in the existing hybrid analysis solutions for fluoresence and surface detector data. This goal has been achieved in a natural way by extending the existing Auger Offline software framework with radio functionality. In this article, we lay out the design, highlights and features of the radio extension implemented in the Auger Offline framework. Its functionality has achieved a high degree of sophistication and offers advanced features such as vectorial reconstruction of the electric field, advanced signal processing algorithms, a transparent and efficient handling of FFTs, a very detailed simulation of detector effects, and the read-in of multiple data formats including data from various radio simulation codes. The source code of this radio functionality can be made available to interested parties on request.Comment: accepted for publication in NIM A, 13 pages, minor corrections to author list and references in v

Search for First Harmonic Modulation in the Right Ascension Distribution of Cosmic Rays Detected at the Pierre Auger Observatory

We present the results of searches for dipolar-type anisotropies in different energy ranges above $2.5\times 10^{17}$ eV with the surface detector array of the Pierre Auger Observatory, reporting on both the phase and the amplitude measurements of the first harmonic modulation in the right-ascension distribution. Upper limits on the amplitudes are obtained, which provide the most stringent bounds at present, being below 2% at 99% $C.L.$ for EeV energies. We also compare our results to those of previous experiments as well as with some theoretical expectations.Comment: 28 pages, 11 figure

Reduced Stability and Increased Dynamics in the Human Proliferating Cell Nuclear Antigen (PCNA) Relative to the Yeast Homolog

Proliferating Cell Nuclear Antigen (PCNA) is an essential factor for DNA replication and repair. PCNA forms a toroidal, ring shaped structure of 90 kDa by the symmetric association of three identical monomers. The ring encircles the DNA and acts as a platform where polymerases and other proteins dock to carry out different DNA metabolic processes. The amino acid sequence of human PCNA is 35% identical to the yeast homolog, and the two proteins have the same 3D crystal structure. In this report, we give evidence that the budding yeast (sc) and human (h) PCNAs have highly similar structures in solution but differ substantially in their stability and dynamics. hPCNA is less resistant to chemical and thermal denaturation and displays lower cooperativity of unfolding as compared to scPCNA. Solvent exchange rates measurements show that the slowest exchanging backbone amides are at the β-sheet, in the structure core, and not at the helices, which line the central channel. However, all the backbone amides of hPCNA exchange fast, becoming undetectable within hours, while the signals from the core amides of scPCNA persist for longer times. The high dynamics of the α-helices, which face the DNA in the PCNA-loaded form, is likely to have functional implications for the sliding of the PCNA ring on the DNA since a large hole with a flexible wall facilitates the establishment of protein-DNA interactions that are transient and easily broken. The increased dynamics of hPCNA relative to scPCNA may allow it to acquire multiple induced conformations upon binding to its substrates enlarging its binding diversity

A complete set of nascent transcription rates for yeast genes

The amount of mRNA in a cell is the result of two opposite reactions: transcription and mRNA degradation. These reactions are governed by kinetics laws, and the most regulated step for many genes is the transcription rate. The transcription rate, which is assumed to be exercised mainly at the RNA polymerase recruitment level, can be calculated using the RNA polymerase densities determined either by run-on or immunoprecipitation using specific antibodies. The yeast Saccharomyces cerevisiae is the ideal model organism to generate a complete set of nascent transcription rates that will prove useful for many gene regulation studies. By combining genomic data from both the GRO (Genomic Run-on) and the RNA pol ChIP-on-chip methods we generated a new, more accurate nascent transcription rate dataset. By comparing this dataset with the indirect ones obtained from the mRNA stabilities and mRNA amount datasets, we are able to obtain biological information about posttranscriptional regulation processes and a genomic snapshot of the location of the active transcriptional machinery. We have obtained nascent transcription rates for 4,670 yeast genes. The median RNA polymerase II density in the genes is 0.078 molecules/kb, which corresponds to an average of 0.096 molecules/gene. Most genes have transcription rates of between 2 and 30 mRNAs/hour and less than 1% of yeast genes have >1 RNA polymerase molecule/gene. Histone and ribosomal protein genes are the highest transcribed groups of genes and other than these exceptions the transcription of genes is an infrequent phenomenon in a yeast cell

Prospects for Cross-correlations of UHECR Events with Astrophysical Sources with Upcoming Space-based Experiments

Ultra-high energy cosmic rays (UHECRs) are the messengers of the most extreme physics in the cosmos; however, efforts to identify their origins have thus far been thwarted by the fact that they don’t point back to their sources. Using statistical studies cross-correlating UHECR arrival directions with astrophysical catalogs, the ground-based Pierre Auger Observatory has reported hints of a correlation with nearby starburst galaxies, as well as lower-significance correlations with other classes of astrophysical sources. Space-based UHECR experiments, such as POEMMA and ZAP, will monitor large interaction volumes on the Earth or the Moon. Within a few years of mission operation time, both missions will achieve unprecedented exposures at energies above 40 EeV across the entire sky. We present studies of the cross-correlation between UHECR event arrival directions and astrophysical catalogs as motivated by expectations for the detector performance for POEMMA and ZAP. We find that both POEMMA and ZAP will achieve 5σ discovery reach for many plausible astrophysical scenarios

Structural and Functional Profiling of the Human Histone Methyltransferase SMYD3

The SET and MYND Domain (SMYD) proteins comprise a unique family of multi-domain SET histone methyltransferases that are implicated in human cancer progression. Here we report an analysis of the crystal structure of the full length human SMYD3 in a complex with an analog of the S-adenosyl methionine (SAM) methyl donor cofactor. The structure revealed an overall compact architecture in which the “split-SET” domain adopts a canonical SET domain fold and closely assembles with a Zn-binding MYND domain and a C-terminal superhelical 9 α-helical bundle similar to that observed for the mouse SMYD1 structure. Together, these structurally interlocked domains impose a highly confined binding pocket for histone substrates, suggesting a regulated mechanism for its enzymatic activity. Our mutational and biochemical analyses confirm regulatory roles of the unique structural elements both inside and outside the core SET domain and establish a previously undetected preference for trimethylation of H4K20

Small Interfering RNA Targeted to IGF-IR Delays Tumor Growth and Induces Proinflammatory Cytokines in a Mouse Breast Cancer Model

Insulin-like growth factor I (IGF-I) and its type I receptor (IGF-IR) play significant roles in tumorigenesis and in immune response. Here, we wanted to know whether an RNA interference approach targeted to IGF-IR could be used for specific antitumor immunostimulation in a breast cancer model. For that, we evaluated short interfering RNA (siRNAs) for inhibition of in vivo tumor growth and immunological stimulation in immunocompetent mice. We designed 2′-O-methyl-modified siRNAs to inhibit expression of IGF-IR in two murine breast cancer cell lines (EMT6, C4HD). Cell transfection of IGF-IR siRNAs decreased proliferation, diminished phosphorylation of downstream signaling pathway proteins, AKT and ERK, and caused a G0/G1 cell cycle block. The IGF-IR silencing also induced secretion of two proinflammatory cytokines, TNF- α and IFN-γ. When we transfected C4HD cells with siRNAs targeting IGF-IR, mammary tumor growth was strongly delayed in syngenic mice. Histology of developing tumors in mice grafted with IGF-IR siRNA treated C4HD cells revealed a low mitotic index, and infiltration of lymphocytes and polymorphonuclear neutrophils, suggesting activation of an antitumor immune response. When we used C4HD cells treated with siRNA as an immunogen, we observed an increase in delayed-type hypersensitivity and the presence of cytotoxic splenocytes against wild-type C4HD cells, indicative of evolving immune response. Our findings show that silencing IGF-IR using synthetic siRNA bearing 2′-O-methyl nucleotides may offer a new clinical approach for treatment of mammary tumors expressing IGF-IR. Interestingly, our work also suggests that crosstalk between IGF-I axis and antitumor immune response can mobilize proinflammatory cytokines