dataesr/scientific-paragraphs-categorization huggingface dataset:
Acknowledgments We are grateful to  M. Durand-Tardif  who first observed the Sha x Mr-0 sterile  F1. J. Drouaud ' advices for long-range amplification and  S. Durand 's remarkably rich database of microsatellite markers were very helpful and appreciated. We gratefully thank  O. Loudet  for the sharing of the neo-Sha population, for numerous stimulating discussions on Arabidopsis natural diversity, and for his constructive comments on an earlier version of the manuscript. We thank  C. Trontin  for sharing her results on the neo-Sha population before publication. P. Touzet,  H. Mireau ,  C. Camilleri  and  G. Pelletier  are gratefully acknowledged for their support, stimulating discussions, and critical comments on earlier versions of the manuscript.
1998; Battesti and Bouveret 2012). We cloned the intact or truncated portions of the rssB coding region as well as each of the anti-adaptor genes in frame with the N-terminal T18 or T25 fragments of cya in compatible plasmids  (Gully and Bouveret 2006). Endpoints for the truncated RssB derivatives were chosen based on the known boundaries of the response regulator domain  (Volz 1995)  and a structure for an RssB-related protein from Pseudomonas aeruginosa (Protein Data Bank [PDB]: 3EQ2) (Fig. 1A ). The resulting recombinant IraP, IraM, IraD, and full-length RssB fusion proteins were functional (Supplemental Fig. S1 ). Fusion proteins with deletions of either the N-terminal or C-terminal domain of RssB were expressed at similar levels (Supplemental Fig. S1C ) and, as expected, were inactive for degradation of RpoS  (Klauck et al.
aeruginosa dimer was created from the RssB structure of P. aeruginosa RssB (PDB: 3EQ2) (I Levchenko, RA Grant, RT Sauer, and TA Baker, unpubl. ) using PYMOL (http://www. pymol. org). One protomer is colored in green; residues mutated during the screen are represented as Corey-Pauling-Koltun (CPK) models. Sequence alignment of RssB E. coli and RssB P. aeruginosa was performed using the ClustalW2 program from the EMBL-EBI Web site (http://www. ebi. ac. uk/ Tools/msa/clustalw2) with default settings. (G) Crystal structure of the RssB N-terminal domain from E. coli. The RssB E. coli N-terminal domain (PDB: 3EOD) was visualized using PYMOL (http://www. pymol. org). Residues on the surface close to the phosphorylation site that were mutated and tested for interaction with IraP are color-coded. Mutations in residues 60, 87, and 88 had no effect on the interaction with IraP; these are colored gray (data not shown).
Several mutant RssB proteins show resistance to multiple anti-adaptors Four mutants isolated as IraP-resistant (P109S, W143R, L214H, and A216T) and two mutants isolated as IraMresistant (L218V and A255V) were chosen as the focus of the work reported here (Fig. 2E ). Although RssB from E. coli has not been crystallized, a P. aeruginosa protein with sequence (24% identity) but not functional homology with E. coli RssB has been crystallized by Baker and coworkers (I Levchenko, RA Grant, RT Sauer, and TA Baker, unpubl. ) (PDB: 3EQ2). In addition, they reported the structure of the N-terminal response regulator domain of E. coli RssB (PDB: 3EOD) (Fig. 2G ). We used the P. aeruginosa structure to show the location of the six mutants used in this study (Fig. 2F ). One, P109S, is in the response regulator domain; another, W143R, lies in the linker.
Mutations selected during the screen for IraP-and IraM-resistant RssB mutants are indicated by an X. The number below each X corresponds to the mutated amino acid based on the E. coli sequence; color-coding of the X corresponds to that used in the graphs in C and D, as well as in the RssB model (F). (F) Localization of mutations on the structure of RssB from P. aeruginosa. The model of the RssB P. aeruginosa dimer was created from the RssB structure of P. aeruginosaRssB (PDB: 3EQ2) (I Levchenko, RA Grant, RT Sauer, and TA Baker, unpubl. ) using PYMOL (http://www. pymol. org). One protomer is colored in green; residues mutated during the screen are represented as Corey-Pauling-Koltun (CPK) models. Sequence alignment of RssB E. coli and RssB P. aeruginosa was performed using the ClustalW2 program from the EMBL-EBI Web site (http://www.
ebi. ac. uk/ Tools/msa/clustalw2) with default settings. (G) Crystal structure of the RssB N-terminal domain from E. coli. The RssB E. coli N-terminal domain (PDB: 3EOD) was visualized using PYMOL (http://www. pymol. org). Residues on the surface close to the phosphorylation site that were mutated and tested for interaction with IraP are color-coded. Mutations in residues 60, 87, and 88 had no effect on the interaction with IraP; these are colored gray (data not shown).
Acknowledgments We thank members of the laboratory,  M. Galperin ,  M. Maurizi , and  D. Dougan , for their comments on the manuscript. We thank  A. Boulanger  for discussions on using the Phos-tag method. This research was supported by the  Intramural Research Program  of the  National Institutes of Health (NIH) ,  National Cancer Institute, Center for Cancer Research. J. M. M. was supported by the  NIH   undergraduate scholarship program.
We used chromatography to isolate a salivary gland protein (Japanin) from Rhipicephalus appendiculatus ticks. Japanin was cloned, and recombinant protein was produced in a baculoviral expression system. We found that Japanin specifically reprogrammes DC responses to a wide variety of stimuli in vitro, radically altering their expression of co-stimulatory and co-inhibitory transmembrane molecules (measured by flow cytometry) and their secretion of pro-inflammatory, anti-inflammatory and T cell polarising cytokines (assessed by Luminex multiplex assays); it also inhibits the differentiation of DC from monocytes. Sequence alignments and enzymatic deglycosylation revealed Japanin to be a 17. 7 kDa, N-glycosylated lipocalin. Using molecular cloning and database searches, we have identified a group of homologous proteins in R.
Multiple rounds of chromatography were then used to isolate the active protein. SGE-3F was first passed through a Q column at pH7. 0, removing many SGE components but leaving the DCmodulatory activity intact. The Q column flow-through was then fractionated using size exclusion chromatography, and a fraction with DC-modulatory activity was further fractionated using HPLC. Activity was detected in a group of consecutive fractions, centred around a single peak on the HPLC trace, from which Edman sequencing generated a 16 residue N-terminal sequence: TPSMPAINTQTLYLAR. We used the above N-terminal sequence to design degenerate primers for polymerase chain reaction (PCR) cloning of a 460 bp sequence from R. appendiculatus salivary gland cDNA (data not shown). This sequence (representing the 39 region of the candidate mRNA) was, in turn, employed to design primers for amplification of the 59 region using 59RACE.
Finally, we performed PCR cloning of the complete coding sequence of the putative DCmodulatory protein, which we named ''Japanin'' (Genbank accession KC412662). Its 531 bp coding sequence encodes a 176 amino acid peptide. Analysis with SignalP 4. 0  [31]  suggests that it is a secretory protein, lacking a transmembrane domain, and comprising a 24 residue cleavable signal peptide followed by a 152 residue mature peptide of predicted 17. 7 kDa molecular weight, the N-terminal sequence of which is consistent with that obtained by Edman degradation. Inspection of the primary amino acid sequence of Japanin indicated that it is a member of the lipocalin family (see below). To facilitate detection and purification of recombinant Japanin, we constructed a polyhistidine-tagged version using PCR. This ''Japanin-his'' DNA sequence (comprising a Kozak consensus sequence for initiation of translation  [32] , the full-length Japanin coding sequence, and a tag sequence encoding a diglycine linker and six histidine residues) was subcloned into bacterial and mammalian expression vectors (pET52b and pCDNA3.
sanguineus, and Japanin like-RA1 and -RA2 (JL-RA1 and JL-RA2) from R. appendiculatus (Genbank accessions KC412663, KC412664, and KC412665; see materials and methods for cloning procedure). Each encodes a 175-177 residue peptide, comprising a 24 residue signal sequence (according to SignalP prediction) and a 151-153 residue mature peptide. Alignment of the predicted mature peptides with Japanin shows that JL-RS is 82% identical and 91% similar to Japanin, JL-RA1 is 50% identical and 74% similar, and JL-RA2 is 54% identical and 76% similar; similarity was calculated using a PAM250 matrix. The high levels of sequence homology between Japanin and the above homologues suggested shared function. To investigate this, we transfected HEK293T cells with pCDNA3. 1 expression constructs encoding either polyhistidine-tagged Japanin, JL-RS, -RA1 or -RA2.
The presence of recombinant protein within the supernatants was confirmed using anti-polyhistidine Western blot (figure  7a ), and the transfectant supernatants were then assessed for their ability to modulate DC maturation, both spontaneously and in response to LPS. Each of the three homologues was found to modulate DC maturation overall (p,0. 05), albeit with some individual differences in the extent of effect on each of these responses, and on CD86 vs. CD274 expression (figures 7b,7c). This suggests that their sequence similarity reflects a shared DCmodulatory function; we are currently evaluating whether or not other modulatory effects are similar to those of Japanin. Finally, in order to search for further Japanin-related sequences in public databases, tblastn searches were performed against the mature peptide sequence of Japanin (see materials and methods).
The NCBI est database returned a cDNA derived from the metastriate tick Dermacentor andersoni with homology [32% identical, 53% similar] to Japanin (Genbank accession EG363153. 1, labelled as DA_E1224_06G04 in figures  8  and 9 ). Furthermore, the Whole-genome shotgun contig database returned regions of Rhipicephalus (Boophilus) microplus genomic DNA whose translations are highly similar to Japanin [40-58% identical, 55-74% similar] (Genbank accessions ADMZ01222530. 1; ADMZ01123695. 1; ADMZ01066468. 1; ADMZ01299354. 1). All the identified homologues conserve Japanin's cysteine residues, which may play a structural role through disulphide bond formation (figure  8 ). Tick lipocalins have conserved intron positions and phase  [49] ; this means that the identification of intron boundaries may be guided by intron-exon structure in addition to the more usual prediction of likely splicing site sequences  [50] [51] [52].
Data shown is from one experiment, representative of three independent experiments using cells from different donors. doi:10. 1371/journal. ppat. 1003450. g005 hard ticks, as well as 3 soft tick proteins with resolved structures. This analysis clearly shows that these molecules form a distinct clade within hard tick lipocalins, grouping in complete isolation from any previously identified proteins or putative proteins and with strong boot-strap support (figure  9 ). Note that for reasons of clarity, only selected proteins are named in figure  9 , and the full tree is provided in Newick (. nwk) format as supplementary data in dataset S1. Japanin is a lipocalin. The mature Japanin sequence was aligned with (A) sequences of mature tick proteins with a resolved lipocalin structure, or (B) with these and additional distantly-related sequences also accepted to be tick lipocalins.
Phylogenetic analysis An initial group of 4 tick lipocalins with published structures (FS-HBP2  [4] , Am182  [86] , Monomine  [86]  and OmCI  [87] ) were aligned using ClustalX  [88] , and this seed alignment used to construct a gap penalty mask. This mask was then employed in the alignment of an additional 242 hard tick lipocalins using ClustalX. Sequences for alignment were taken from: (i) the table provided as supplementary data by Francischetti and colleagues  [89] , from which all complete sequences identified as hard tick lipocalins, were used, with the exception of those described as group VIII, which we do not believe to be lipocalins (based on the absence of conserved sequence features); (ii) LIR2 and LIR7  [90] ; (iii) Ir-LBP  [91] ; (iv) the sequences described in this paper. Sequences are named according to their published abbreviation, Genbank accession number, or as referred to by Francischetti and colleagues.
All positions with less than 90% site coverage were eliminated. That is, no more than 10% alignment gaps, missing data, or ambiguous bases were allowed at any position. There were a total of 112 positions in the final dataset. An alternative analysis where all positions with less than 95% site coverage were supported the conclusion that Japanin-like proteins form a clade, as did an analysis using the neighbour-joining method (with evolutionary distances computed using the JTT matrix-based method).
Acknowledgments We thank  Daniel Lunn  for statistical analysis; Jan Digby and  Gareth Williams  for Luminex data acquisition and analysis;  Philipp Becker  for trial experiments;  Wilson Tang  for homologue protein expression;  Julie Schulthess  and  Carolina Arancibia  for assistance with PBMC subset staining and for supplying antibodies; and  Susan Lea  for comments on the manuscript.
Software Statistical analyses were performed with R  [94] , using the lme4  [95]  and ordinal  [96]  packages for modelling, and the languageR package  [97]  for MCMC sampling. Line and bar charts were produced with R, using the ggplot2  [98] , plotrix  [99]  and Cairo  [100]  packages. Flow cytometry data was collected using FACSDiva (BD Biosciences), and analysed and plotted with FlowJo (Tree Star). Sequence alignments were viewed and edited using UGENE  [101] , and formatted for publication with Jalview. Phylogenetic analyses were conducted using MEGA version 5. 1  [102] , and trees were formatted for publication with FigTree 1. 4 (http://tree. bio. ed. ac. uk/software/figtree/).
While the debate about climate modelling is important, it needs to be informed by empirical evidence. Before reviewing quantitative analyses, consider Fig. 1  that shows examples of temperatures from two scales within the weather regime (0. 067 s, 1 h) and at two lower resolutions (top curves, 20 days and 1 century). Other atmospheric fields  (wind, humidity, precipitation, etc. )   To make the comparison as fair as possible, in each case, the sample is 720 points long and each series has its mean removed and is normalized by its standard deviation (0. 35, 4. 49, 2. 59, 1. 39 K, respectively), the three upper series have been displaced in the vertical by four units for clarity. The resolutions are 0. 067 s, 1 h, 20 days and 1 century, respectively, the data are from 4 m above the top of the roof of the Rutherford physics building (Montreal, Quebec), a weather station in Lander, Wyoming, the 20th century reanalysis and the Vostok Antarctic station, respectively.
Relevant MatLab and Mathematica software are available at http://www. physics. mcgill. ca/~gang/software/index. html. In a scaling regime, S q ( t) is a power law; S q ( t) ≈ t ξ (q) , where the exponent ξ(q) = q H -K(q) and K(q) characterizes the scaling intermittency (with K(1) = 0). In the macroweather regime K(2) is small (≈ 0. 01-0. 03), so that the RMS (root mean square) variation S 2 ( t) 1/2 (denoted simply S( t) below) has the exponent ξ (2)/2 ≈ ξ (1) = H. In the climate regime the intermittency correction is a bit larger  (Schmitt et al. , 1995)  (≈ 0. 12) but the error in using this approximation (≈ 0. 06) will be neglected.
03 W kg -1 and τ w,mars ≈ 1. 5 days. This has recently been confirmed using in situ temperature and wind data from the Viking Mars lander (manuscript in preparation); there are thus three examples of weathermacroweather transitions. Δ Δ Fig. 4. Control runs versus preindustrial multiproxies: a comparison of the RMS Haar structure functions (S( t)) for temperatures from the multiproxies Fig. 3  (resolution 1 yr, black, 1500-1900) and GCM control runs (brown dashed, IPSL, EFS monthly), annual resolution GISS-E2-R, (thick brown, continuous) and the FIF stochastic model (thin brown). With the exception of the GISS-E2-R (land, Northern Hemisphere), the data are averaged over the entire globe. The IPSL curve is from a 500 yr control run, the EFS is from a 3000 yr control run; the "bump" at 2-4 yr is a broad quasi periodic model artefact.
The last millennium simulations: the climate or macroweather? If control runs produce only fluctuations decreasing with scale (macroweather), what about forced runs with more realistic low frequency variability? To answer this question, we must systematically compare the model and empirical fluctuations as functions of timescale t. However, first we must consider a complication: the effect of anthropogenic forcings over the industrial period. We saw earlier (Fig. 3 ) that for scales t ≈ > 10 yr there was a large difference between the industrial (1880-present) surface series and preindustrial (1500-1900) multiproxies -presumably a consequence of the fact that the recent period variability of the global temperatures is dominated by anthropogenic effects. Figure  5  shows the analysis of eight GISS-E2-R last millennium simulations (Northern Hemisphere, land only) over the period 1880-2005 (CMIP5 database, see also  Schmidt et al.
2 mJy kpc 2 of only 3. 0(±0. 7) × 10 4 MSPs. We have also simulated pulsars with periods shorter than any known MSP, and estimate the maximum number of sub-MSPs in the Galaxy to be 7. 8(±5. 0) × 10 4 pulsars at L = 0. 1 mJy kpc 2. In addition, we estimate that the high and low latitude parts of the southern HTRU survey will detect 68 and 42 MSPs respectively, including 78 new discoveries. Pulsar luminosity, and hence flux density, is an important input parameter in the model. Some of the published flux densities for the pulsars in our sample do not agree with the observed flux densities from our data set, and we have instead calculated average luminosities from archival data from the Parkes Telescope. We found many luminosities to be very different than their catalogue values, leading to very different population estimates.
As For this analysis we have used 50 previously known and newly discovered recycled pulsars in the HTRU medlat survey area not associated with globular clusters. We define recycled pulsars as having a period P < 0. 070 s and a spin down rate Ṗ < 10 -17 s/s. All pulsars in our sample are listed in Table  2  together with some of their properties and a note of in which 20-cm pulsar surveys they have been detected. Three of the newly discovered pulsars from the HTRU medlat survey were confirmed only shortly before this work started. Timing observations of these pulsars are carried out at Jodrell Bank Observatory and due to a lack of data from the Parkes Telescope at the time, these pulsars are not included in this analysis. To make sure the numbers of detected pulsars from our simulation in each survey was correct, archival data from the Parkes Multibeam survey (PM) and the Swinburne Intermediate Latitude survey (SIL) were searched for the sample pulsars.
Analysis We have used the program psrevolve 5  to generate pulsars randomly spread out in the Galaxy and check if they are detectable in a number of radio pulsar surveys. To start with, we simulated 150,000 pulsars with periods, pulse widths and luminosities identical to each of the input pulsars in Table  2. These were spread out in the Galaxy assuming a Gaussian radial distribution with radial scale length R = 4. 5 kpc and z-values from a Gaussian distribution with a root mean square height z = 500 pc. The scale height was later varied to investigate the z-height of the underlying population. A snapshot approach was used, in which it is checked whether a pulsar is detectable by different pulsar surveys directly where it is placed by the simulation. For each of the surveys used in this analysis, we have created a database of the coordinates for each observation.
Each pointing in the hilat survey is integrated for 270 s, which is half the integration time of the medlat survey observations. In the first survey paper of the HTRU survey, Keith et al. (  2010 ) simulated the estimated detection and discovery numbers for the each of sub-surveys, using the pulsar population model de-scribed by  Lorimer et al. (2006)  and the PSRPOP 6  software. The main difference between that simulation and the analysis described here is that Keith et al. assumed a luminosity distribution as a power law with index -0. 59 with minimum luminosity Lmin = 0. 1 mJy kpc 2. The Keith et al. simulation returned a similar number of medlat detections with 48 detections (and 28 discoveries) compared to the actual numbers of MSPs detected in the survey (51 detections, 26 discoveries).
ACKNOWLEDGEMENTS The work presented in this paper was conducted as part of  L. Levin 's PhD thesis at Swinburne University and was originally written as a thesis chapter. The version presented here has been slightly modified. This work is supported by the  ARC Centre of Excellence for All-sky Astrophysics (CAASTRO). The Parkes Observatory is part of the  Australia Telescope , which is funded by the  Commonwealth of Australia  for operation as a National Facility managed by  CSIRO. This paper includes archived data obtained through the  Australia Telescope Online Archive  and the  CSIRO Data Access Portal  (http://data. csiro. au). The authors would like to thank the referee for helpful comments and suggestions.
The quantified signals were normalized using the global Lowess (Locally Weighted Scatterplot Smoothing) regression algorithm, which minimizes the intensity-dependent differences between the dyes. The expression matrix contains normalized Hy3/Hy5 ratios (log2 transformed) from all hybridizations. The subset of differentially expressed miRNAs was used to construct an unsupervised hierarchical clustering of the different samples (Exiqon, A/S). For in silico data analysis, we used the GOmir program (http://www. bioacademy. gr/bioinformatics/projects/GOmir/), which integrates miRNA target prediction and functional analysis by combining the predicted target genes from the TargetScan (http:// www. targetscan. org/), miRanda (http://www. microrna. org/ microrna/home. do), RNAhybrid (http://bibiserv.
Fibroblast Colony-Forming Unit Assay The clonogenic potential of SS-AF-MSCs expressing miR-21 or sh-Sox2 was evaluated by performing a fibroblast colony-forming unit (CFU-F) assay. Specifically, transduced SS-AF-MSCs expressing miR-21, EF1, sh-Sox2, or sh-control were plated at three clonal densities (70, 140, and 280 cells) in 60-mm plates for 14 days. The CFU-Fs were quantified by Giemsa staining. The number of CFU-Fs was estimated per 100 MSCs initially plated based on a linear regression analysis of the three different initial cell concentrations. The data are presented as the mean 6 SEM of at least three independent experiments.
Cell Cycle Analysis SS-AF-MSCs expressing miR-21 or EF1 were stained with propidium iodide. The cells were fixed in ice-cold 70% ethanol for at least 16 hours at 4°C. After fixation, the cells were incubated with 2 mg/ml RNase A (Sigma-Aldrich) at 37°C for 40 minutes. Then, 50 mg/ml propidium iodide (Sigma-Aldrich) was added, and the cells were analyzed by flow cytometry. The nuclear debris, and overlapping nuclei were gated out. The data are presented as the mean 6 SEM of at least three independent experiments. Senescence-associated b-galactosidase (SA b-gal) staining was performed using a cellular senescence assay kit (Merck KGaA). Briefly, 5 3 10 4 transduced SS-AF-MSCs expressing miR-21 or EF1 were fixed in 13 fixing solution and stained for SA b-gal activity overnight. The positive (blue) cells were counted and were expressed relative to the number of positive cells in SS-AF-MSCs expressing EF1.