Role of the Bombyx mori R2 element N-terminal domain in the target-primed reverse transcription (TPRT) reaction

R2 is a site-specific non-long terminal repeat (non-LTR) retrotransposon encoding a single polypeptide with reverse transcriptase, DNA endonuclease and nucleic acid-binding domains. The current model of R2 retrotransposition involves an ordered series of cleavage and polymerization steps carried out by at least two R2 protein subunits, one bound upstream and one bound downstream of the integration site. The role in the retrotransposition reaction of two conserved DNA-binding motifs, a C(2)H(2) zinc finger (ZF) and a Myb motif, located within the N-terminal domain of the protein are explored in this report. These motifs do not appear to play a role in RT or the ability of the protein to bind the R2 RNA transcript. Methylation and missing nucleoside interference-based DNA footprints using polypeptides to the N-terminal domain suggest the ZF and Myb motifs bind to regions −3 to −1 and +10 to +15 with reference to the insertion site. Mutations in these DNA sites or of the N-terminal protein domain blocked binding and the activity of the downstream subunit. Mutations of the protein domain also affected binding of the upstream subunit but not its function, suggesting the primary path to DNA target recognition by R2 involves both upstream and downstream subunits

Effects of elevated [CO2 ] on maize defence against mycotoxigenic Fusarium verticillioides.

Maize is by quantity the most important C4 cereal crop; however, future climate changes are expected to increase maize susceptibility to mycotoxigenic fungal pathogens and reduce productivity. While rising atmospheric [CO2 ] is a driving force behind the warmer temperatures and drought, which aggravate fungal disease and mycotoxin accumulation, our understanding of how elevated [CO2 ] will effect maize defences against such pathogens is limited. Here we report that elevated [CO2 ] increases maize susceptibility to Fusarium verticillioides proliferation, while mycotoxin levels are unaltered. Fumonisin production is not proportional to the increase in F. verticillioides biomass, and the amount of fumonisin produced per unit pathogen is reduced at elevated [CO2 ]. Following F. verticillioides stalk inoculation, the accumulation of sugars, free fatty acids, lipoxygenase (LOX) transcripts, phytohormones and downstream phytoalexins is dampened in maize grown at elevated [CO2 ]. The attenuation of maize 13-LOXs and jasmonic acid production correlates with reduced terpenoid phytoalexins and increased susceptibility. Furthermore, the attenuated induction of 9-LOXs, which have been suggested to stimulate mycotoxin biosynthesis, is consistent with reduced fumonisin per unit fungal biomass at elevated [CO2 ]. Our findings suggest that elevated [CO2 ] will compromise maize LOX-dependent signalling, which will influence the interactions between maize and mycotoxigenic fungi

Genome wide screens in yeast to identify potential binding sites and target genes of DNA-binding proteins

Knowledge of all binding sites for transcriptional activators and repressors is essential for computationally aided identification of transcriptional networks. The techniques developed for defining the binding sites of transcription factors tend to be cumbersome and not adaptable to high throughput. We refined a versatile yeast strategy to rapidly and efficiently identify genomic targets of DNA-binding proteins. Yeast expressing a transcription factor is mated to yeast containing a library of genomic fragments cloned upstream of the reporter gene URA3. DNA fragments with target-binding sites are identified by growth of yeast clones in media lacking uracil. The experimental approach was validated with the tumor suppressor protein p53 and the forkhead protein FoxI1 using genomic libraries for zebrafish and mouse generated by shotgun cloning of short genomic fragments. Computational analysis of the genomic fragments recapitulated the published consensus-binding site for each protein. Identified fragments were mapped to identify the genomic context of each binding site. Our yeast screening strategy, combined with bioinformatics approaches, will allow both detailed and high-throughput characterization of transcription factors, scalable to the analysis of all putative DNA-binding proteins

Planetary Candidates Observed by Kepler. VIII. A Fully Automated Catalog With Measured Completeness and Reliability Based on Data Release 25

We present the Kepler Object of Interest (KOI) catalog of transiting exoplanets based on searching four years of Kepler time series photometry (Data Release 25, Q1-Q17). The catalog contains 8054 KOIs of which 4034 are planet candidates with periods between 0.25 and 632 days. Of these candidates, 219 are new and include two in multi-planet systems (KOI-82.06 and KOI-2926.05), and ten high-reliability, terrestrial-size, habitable zone candidates. This catalog was created using a tool called the Robovetter which automatically vets the DR25 Threshold Crossing Events (TCEs, Twicken et al. 2016). The Robovetter also vetted simulated data sets and measured how well it was able to separate TCEs caused by noise from those caused by low signal-to-noise transits. We discusses the Robovetter and the metrics it uses to sort TCEs. For orbital periods less than 100 days the Robovetter completeness (the fraction of simulated transits that are determined to be planet candidates) across all observed stars is greater than 85%. For the same period range, the catalog reliability (the fraction of candidates that are not due to instrumental or stellar noise) is greater than 98%. However, for low signal-to-noise candidates between 200 and 500 days around FGK dwarf stars, the Robovetter is 76.7% complete and the catalog is 50.5% reliable. The KOI catalog, the transit fits and all of the simulated data used to characterize this catalog are available at the NASA Exoplanet Archive.Comment: 61 pages, 23 Figures, 9 Tables, Accepted to The Astrophysical Journal Supplement Serie

Isoenergetic penta- and hexanucleotide microarray probing and chemical mapping provide a secondary structure model for an RNA element orchestrating R2 retrotransposon protein function

LNA (locked nucleic acids, i.e. oligonucleotides with a methyl bridge between the 2′ oxygen and 4′ carbon of ribose) and 2,6-diaminopurine were incorporated into 2′-O-methyl RNA pentamer and hexamer probes to make a microarray that binds unpaired RNA approximately isoenergetically. That is, binding is roughly independent of target sequence if target is unfolded. The isoenergetic binding and short probe length simplify interpretation of binding to a structured RNA to provide insight into target RNA secondary structure. Microarray binding and chemical mapping were used to probe the secondary structure of a 323 nt segment of the 5′ coding region of the R2 retrotransposon from Bombyx mori (R2Bm 5′ RNA). This R2Bm 5′ RNA orchestrates functioning of the R2 protein responsible for cleaving the second strand of DNA during insertion of the R2 sequence into the genome. The experimental results were used as constraints in a free energy minimization algorithm to provide an initial model for the secondary structure of the R2Bm 5′ RNA

Planetary Candidates Observed by Kepler, III: Analysis of the First 16 Months of Data

New transiting planet candidates are identified in sixteen months (May 2009 - September 2010) of data from the Kepler spacecraft. Nearly five thousand periodic transit-like signals are vetted against astrophysical and instrumental false positives yielding 1,091 viable new planet candidates, bringing the total count up to over 2,300. Improved vetting metrics are employed, contributing to higher catalog reliability. Most notable is the noise-weighted robust averaging of multi-quarter photo-center offsets derived from difference image analysis which identifies likely background eclipsing binaries. Twenty-two months of photometry are used for the purpose of characterizing each of the new candidates. Ephemerides (transit epoch, T_0, and orbital period, P) are tabulated as well as the products of light curve modeling: reduced radius (Rp/R*), reduced semi-major axis (d/R*), and impact parameter (b). The largest fractional increases are seen for the smallest planet candidates (197% for candidates smaller than 2Re compared to 52% for candidates larger than 2Re) and those at longer orbital periods (123% for candidates outside of 50-day orbits versus 85% for candidates inside of 50-day orbits). The gains are larger than expected from increasing the observing window from thirteen months (Quarter 1-- Quarter 5) to sixteen months (Quarter 1 -- Quarter 6). This demonstrates the benefit of continued development of pipeline analysis software. The fraction of all host stars with multiple candidates has grown from 17% to 20%, and the paucity of short-period giant planets in multiple systems is still evident. The progression toward smaller planets at longer orbital periods with each new catalog release suggests that Earth-size planets in the Habitable Zone are forthcoming if, indeed, such planets are abundant.Comment: Submitted to ApJS. Machine-readable tables are available at http://kepler.nasa.gov, http://archive.stsci.edu/kepler/results.html, and the NASA Exoplanet Archiv

The Occurrence of Rocky Habitable-zone Planets around Solar-like Stars from Kepler Data

We present the occurrence rates for rocky planets in the habitable zones (HZs) of main-sequence dwarf stars based on the Kepler DR25 planet candidate catalog and Gaia-based stellar properties. We provide the first analysis in terms of star-dependent instellation flux, which allows us to track HZ planets. We define η⊕ as the HZ occurrence of planets with radii between 0.5 and 1.5 R⊕ orbiting stars with effective temperatures between 4800 and 6300 K. We find that η⊕ for the conservative HZ is between 0.37^(+0.48)_(−0.21) (errors reflect 68% credible intervals) and 0.60^(+0.90)_(−0.36) planets per star, while the optimistic HZ occurrence is between 0.58^(+0.73)_(−0.33) and 0.88^(+1.28)_(−0.51) planets per star. These bounds reflect two extreme assumptions about the extrapolation of completeness beyond orbital periods where DR25 completeness data are available. The large uncertainties are due to the small number of detected small HZ planets. We find similar occurrence rates between using Poisson likelihood Bayesian analysis and using Approximate Bayesian Computation. Our results are corrected for catalog completeness and reliability. Both completeness and the planet occurrence rate are dependent on stellar effective temperature. We also present occurrence rates for various stellar populations and planet size ranges. We estimate with 95% confidence that, on average, the nearest HZ planet around G and K dwarfs is ~6 pc away and there are ~4 HZ rocky planets around G and K dwarfs within 10 pc of the Sun

The Occurrence of Rocky Habitable Zone Planets Around Solar-Like Stars from Kepler Data

We present occurrence rates for rocky planets in the habitable zones (HZ) of main-sequence dwarf stars based on the Kepler DR25 planet candidate catalog and Gaia-based stellar properties. We provide the first analysis in terms of star-dependent instellation flux, which allows us to track HZ planets. We define $\eta_\oplus$ as the HZ occurrence of planets with radius between 0.5 and 1.5 $R_\oplus$ orbiting stars with effective temperatures between 4800 K and 6300 K. We find that $\eta_\oplus$ for the conservative HZ is between $0.37^{+0.48}_{-0.21}$ (errors reflect 68\% credible intervals) and $0.60^{+0.90}_{-0.36}$ planets per star, while the optimistic HZ occurrence is between $0.58^{+0.73}_{-0.33}$ and $0.88^{+1.28}_{-0.51}$ planets per star. These bounds reflect two extreme assumptions about the extrapolation of completeness beyond orbital periods where DR25 completeness data are available. The large uncertainties are due to the small number of detected small HZ planets. We find similar occurrence rates using both a Poisson likelihood Bayesian analysis and Approximate Bayesian Computation. Our results are corrected for catalog completeness and reliability. Both completeness and the planet occurrence rate are dependent on stellar effective temperature. We also present occurrence rates for various stellar populations and planet size ranges. We estimate with $95\%$ confidence that, on average, the nearest HZ planet around G and K dwarfs is about 6 pc away, and there are about 4 HZ rocky planets around G and K dwarfs within 10 pc of the Sun.Comment: To appear in The Astronomical Journa

The FLUXNET2015 dataset and the ONEFlux processing pipeline for eddy covariance data

The FLUXNET2015 dataset provides ecosystem-scale data on CO2, water, and energy exchange between the biosphere and the atmosphere, and other meteorological and biological measurements, from 212 sites around the globe (over 1500 site-years, up to and including year 2014). These sites, independently managed and operated, voluntarily contributed their data to create global datasets. Data were quality controlled and processed using uniform methods, to improve consistency and intercomparability across sites. The dataset is already being used in a number of applications, including ecophysiology studies, remote sensing studies, and development of ecosystem and Earth system models. FLUXNET2015 includes derived-data products, such as gap-filled time series, ecosystem respiration and photosynthetic uptake estimates, estimation of uncertainties, and metadata about the measurements, presented for the first time in this paper. In addition, 206 of these sites are for the first time distributed under a Creative Commons (CC-BY 4.0) license. This paper details this enhanced dataset and the processing methods, now made available as open-source codes, making the dataset more accessible, transparent, and reproducible.Peer reviewe