The SNF2-Family Member Fun30 Promotes Gene Silencing in Heterochromatic Loci

Chromatin regulates many key processes in the nucleus by controlling access to the underlying DNA. SNF2-like factors are ATP-driven enzymes that play key roles in the dynamics of chromatin by remodelling nucleosomes and other nucleoprotein complexes. Even simple eukaryotes such as yeast contain members of several subfamilies of SNF2-like factors. The FUN30/ETL1 subfamily of SNF2 remodellers is conserved from yeasts to humans, but is poorly characterized. We show that the deletion of FUN30 leads to sensitivity to the topoisomerase I poison camptothecin and to severe cell cycle progression defects when the Orc5 subunit is mutated. We demonstrate a role of FUN30 in promoting silencing in the heterochromatin-like mating type locus HMR, telomeres and the rDNA repeats. Chromatin immunoprecipitation experiments demonstrate that Fun30 binds at the boundary element of the silent HMR and within the silent HMR. Mapping of nucleosomes in vivo using micrococcal nuclease demonstrates that deletion of FUN30 leads to changes of the chromatin structure at the boundary element. A point mutation in the ATP-binding site abrogates the silencing function of Fun30 as well as its toxicity upon overexpression, indicating that the ATPase activity is essential for these roles of Fun30. We identify by amino acid sequence analysis a putative CUE motif as a feature of FUN30/ETL1 factors and show that this motif assists Fun30 activity. Our work suggests that Fun30 is directly involved in silencing by regulating the chromatin structure within or around silent loci

Enduro Campers

The Enduro Campers senior project team (Formerly known as Lobo Campers senior project team) was recruited by Mr. Leif Stein to assist in the design, manufacturing, and testing of the materials and components of his company’s expedition box camper. The two main focuses of the project, divided into two sub-teams, were on the testing and manufacturing process of the materials used in the camper, and on the design and analysis of the camper’s access door, step box, and other subsystems. The focus of the materials analysis and manufacturing team was based around the design parameter to use composite sandwich panels for the majority of the camper structure as set by Mr. Stein. This made the manufacturing and analysis of the camper much more complex than if a homogeneous material such as aluminum was used for the structure’s body. This increase in complexity is because composites are more difficult than homogenous materials to manufacture, test, and analyze. Because of this, the company needed a standardized physical testing procedure as well as an optimized bonding process for the sandwich materials. In order to choose the combination of materials for the sandwich panel construction, the project required a material analysis through physical testing and development of an FEA material model. The program ABAQUS was used to model the sandwich panel and can be applied to future analyses that Mr. Stein will need for larger, specific sections of the camper’s structure under different loading cases. The sub-component design & analysis team delivered a design for the door and step box of the camper that allows for maximum accessibility while maintaining the camper’s structural integrity while sealing the cabin from the elements. This design is also aesthetically pleasing, as these are the main visible components on the expedition box product. The design derived by our team was based on input from Mr. Stein, as well as the findings from the materials research team through analysis and physical testing. The manufacturing process of the step box was started and plans were handed to Mr. Stein for completion. This final design report goes over the project’s scope of work, concept design development, final design choices, manufacturing, design verification, project management, and the conclusions and recommendations the team has for Mr. Stein and his developing company. The report also includes recommendations for future analysis that can be helpful to better understanding the sandwich material used in the construction of the camper

Selection of chromosomal DNA libraries using a multiplex CRISPR system.

The directed evolution of biomolecules to improve or change their activity is central to many engineering and synthetic biology efforts. However, selecting improved variants from gene libraries in living cells requires plasmid expression systems that suffer from variable copy number effects, or the use of complex marker-dependent chromosomal integration strategies. We developed quantitative gene assembly and DNA library insertion into the Saccharomyces cerevisiae genome by optimizing an efficient single-step and marker-free genome editing system using CRISPR-Cas9. With this Multiplex CRISPR (CRISPRm) system, we selected an improved cellobiose utilization pathway in diploid yeast in a single round of mutagenesis and selection, which increased cellobiose fermentation rates by over 10-fold. Mutations recovered in the best cellodextrin transporters reveal synergy between substrate binding and transporter dynamics, and demonstrate the power of CRISPRm to accelerate selection experiments and discoveries of the molecular determinants that enhance biomolecule function

Metabolic scaling in modular animals

Metabolic scaling is the relationship between organismal metabolic rate and body mass. Understanding the patterns and causes of metabolic scaling provides a powerful foundation for predicting biological processes at the level of individuals, populations, communities, and ecosystems. Despite intense interest in, and debate on, the mechanistic basis of metabolic scaling, relatively little attention has been paid to metabolic scaling in clonal animals with modular construction, such as colonial cnidarians, bryozoans, and colonial ascidians. Unlike unitary animals, modular animals are structural individuals subdivided into repeated morphological units, or modules, each able to acquire, process, and share resources. A modular design allows flexibility in organism size and shape with consequences for metabolic scaling. Furthermore, with careful consideration of the biology of modular animals, the size and shape of individual colonies can be experimentally manipulated to test competing theories pertaining to metabolic scaling. Here, we review metabolic scaling in modular animals and find that a wide range of scaling exponents, rather than a single value, has been reported for a variety of modular animals. We identify factors influencing variation in intraspecific scaling in this group that relate to the general observation that not all modules within a colony are identical. We highlight current gaps in our understanding of metabolic scaling in modular animals, and suggest future research directions, such as manipulating metabolic states and comparisons among species that differ in extent of module integration

Post-Newtonian expansion of gravitational waves from a particle in circular orbits around a rotating black hole: Up to O(v8)O(v^8) beyond the quadrupole formula

Extending a method developed by Sasaki in the Schwarzschild case and by Shibata, Sasaki, Tagoshi, and Tanaka in the Kerr case, we calculate the post-Newtonian expansion of the gravitational wave luminosities from a test particle in circular orbit around a rotating black hole up to $O(v^8)$ beyond the quadrupole formula. The orbit of a test particle is restricted on the equatorial plane. We find that spin dependent terms appear in each post-Newtonian order, and that at $O(v^6)$ they have a significant effect on the orbital phase evolution of coalescing compact binaries. By comparing the post-Newtonian formula of the luminosity with numerical results we find that, for $30M\lesssim r \lesssim 100M$, the spin dependent terms at $O(v^6)$ and $O(v^7)$ improve the accuracy of the post-Newtonian formula significantly, but those at $O(v^8)$ do not improve.Comment: 27 pages, revtex, 6 figures, submitted to Physical Review

Spin effects in gravitational radiation backreaction III. Compact binaries with two spinning components

The secular evolution of a spinning, massive binary system in eccentric orbit is analyzed, expanding and generalizing our previous treatments of the Lense-Thirring motion and the one-spin limit. The spin-orbit and spin-spin effects up to the 3/2 post-Newtonian order are considered, both in the equations of motion and in the radiative losses. The description of the orbit in terms of the true anomaly parametrization provides a simple averaging technique, based on the residue theorem, over eccentric orbits. The evolution equations of the angle variables characterizing the relative orientation of the spin and orbital angular momenta reveal a speed-up effect due to the eccentricity. The dissipative evolutions of the relevant dynamical and angular variables is presented in the form of a closed system of differential equations.Comment: 10 pages, 1 figur

Comparative metagenomics reveals a diverse range of antimicrobial resistance genes in effluents entering a river catchment.

The aquatic environment has been implicated as a reservoir for antimicrobial resistance genes (ARGs). In order to identify sources that are contributing to these gene reservoirs, it is crucial to assess effluents that are entering the aquatic environment. Here we describe a metagenomic assessment for two types of effluent entering a river catchment. We investigated the diversity and abundance of resistance genes, mobile genetic elements (MGEs) and pathogenic bacteria. Findings were normalised to a background sample of river source water. Our results show that effluent contributed an array of genes to the river catchment, the most abundant being tetracycline resistance genes tetC and tetW from farm effluents and the sulfonamide resistance gene sul2 from wastewater treatment plant (WWTP) effluents. In nine separate samples taken across 3 years, we found 53 different genes conferring resistance to seven classes of antimicrobial. Compared to the background sample taken up river from effluent entry, the average abundance of genes was three times greater in the farm effluent and two times greater in the WWTP effluent. We conclude that effluents disperse ARGs, MGEs and pathogenic bacteria within a river catchment, thereby contributing to environmental reservoirs of ARGs.This research was funded by the Centre for Environment, Fisheries and Aquaculture Science, GlaxoSmithKline and the Biotechnology and Biological Sciences Research Council. Metagenomic sequencing was performed by the Sequencing Service and Wellcome Trust Biomedical Informatics Hub, Exeter (UK).This is the author accepted manuscript. It is currently under an indefinite embargo pending publication by IWA Publishing

Black Hole Spectroscopy: Testing General Relativity through Gravitational Wave Observations

Assuming that general relativity is the correct theory of gravity in the strong field limit, can gravitational wave observations distinguish between black hole and other compact object sources? Alternatively, can gravitational wave observations provide a test of one of the fundamental predictions of general relativity? Here we describe a definitive test of the hypothesis that observations of damped, sinusoidal gravitational waves originated from a black hole or, alternatively, that nature respects the general relativistic no-hair theorem. For astrophysical black holes, which have a negligible charge-to-mass ratio, the black hole quasi-normal mode spectrum is characterized entirely by the black hole mass and angular momentum and is unique to black holes. In a different theory of gravity, or if the observed radiation arises from a different source (e.g., a neutron star, strange matter or boson star), the spectrum will be inconsistent with that predicted for general relativistic black holes. We give a statistical characterization of the consistency between the noisy observation and the theoretical predictions of general relativity, together with a numerical example.Comment: 19 pages, 7 figure

Gravitational waves from coalescing binaries and Doppler experiments

Doppler tracking of interplanetary spacecraft provides the only method presently available for broad-band searches of low frequency gravitational waves. The instruments have a peak sensitivity around the reciprocal of the round-trip light-time T of the radio link connecting the Earth to the space-probe and therefore are particularly suitable to search for coalescing binaries containing massive black holes in galactic nuclei. A number of Doppler experiments -- the most recent involving the probes ULYSSES, GALILEO and MARS OBSERVER -- have been carried out so far; moreover, in 2002-2004 the CASSINI spacecraft will perform three 40 days data acquisition runs with expected sensitivity about twenty times better than that achieved so far. Central aims of this paper are: (i) to explore, as a function of the relevant instrumental and astrophysical parameters, the Doppler output produced by in-spiral signals -- sinusoids of increasing frequency and amplitude (the so-called chirp); (ii) to identify the most important parameter regions where to concentrate intense and dedicated data analysis; (iii) to analyze the all-sky and all-frequency sensitivity of the CASSINI's experiments, with particular emphasis on possible astrophysical targets, such as our Galactic Centre and the Virgo Cluster.Comment: 52 pages, LaTeX, 19 Postscript Figures, submitted to Phys. Rev.

The Confrontation between General Relativity and Experiment

The status of experimental tests of general relativity and of theoretical frameworks for analysing them are reviewed. Einstein's equivalence principle (EEP) is well supported by experiments such as the E\"otv\"os experiment, tests of special relativity, and the gravitational redshift experiment. Future tests of EEP and of the inverse square law will search for new interactions arising from unification or quantum gravity. Tests of general relativity at the post-Newtonian level have reached high precision, including the light deflection, the Shapiro time delay, the perihelion advance of Mercury, and the Nordtvedt effect in lunar motion. Gravitational wave damping has been detected to half a percent using the binary pulsar, and new binary pulsar systems may yield further improvements. When direct observation of gravitational radiation from astrophysical sources begins, new tests of general relativity will be possible.Comment: 103 pages, 10 figures, accepted for publication in Living Reviews in Relativit