dataesr/scientific-paragraphs-categorization huggingface dataset:
6  The two sets of generated data were affected differently as shown by the average CV as a function of reporter ion intensity. Oxidized cysteine percentages are calculated based on the oxidized cysteine reporter ion which is much prone to be affected by contamination by comparison with the more intense total cysteine reporter ion. The CVs for the oxidized fraction decrease with the reporter's intensity, stabilizing under 30%, our limit of acceptance for a quantitative value  Another limitation of the method is that the calculation of the expression profiles is limited to cysteinecontaining peptides. The higher the number of peptides used for the quantification, the more robust the quantitative information is. When performing protein quantification using several peptides, outliers (often peptides affected by multiple PTMs) are easily detected and eliminated.
Twenty-seven serpins belonging to clade A, B, C, D, E, F, G, H and I serpins are currently referenced in chicken genome databases. Phylogenetic analysis of chicken serpins revealed that ovalbumin (Serpinb14) and its paralogs ovalbumin-related protein Y (Serpinb14b) and ovalbumin-related protein X (Serpinb14c) are found in bird species. These clade B serpins are specifically expressed in reproductive tissues and exported in the egg where they constitute major protein components. These data suggest that these three paralogs have probably appeared in birds to face new environments and ensure the extra-uterine development of an embryo in a shell egg. Twelve other serpins have been identified in the newly produced egg, some of them having a specific distribution in the respective egg structures (eggshell, egg white, vitelline membrane and egg yolk).
Evolution and distribution of egg serpins Systematic analysis of National Center for Biotechnology Information and chicken Ensembl databases for identifying serpins in chicken species revealed the presence of 27 members of this family (Table  1 , Fig. 1 ). Among these, 16 serpins are still predicted and have been referenced in databases by automated computational analysis of genomic chicken sequences annotated using gene prediction methods, which sometimes can lead to discrepancies between gene and protein names (as an example, Serpinb2 gene corresponds to a predicted Serpinb10 protein, Table  1 ). However, the relevance of 5 of them have been recently validated, as serpins a8, c1, d1 and f1 were unambiguously identified in the egg (see §1. 2). Thus, 11 of the serpins identified to date from genome analysis, need further validation and the information available about their physiological functions is therefore very partial, even inexistent.
Surprisingly enough, Serpine1 was not identified in this analysis. To investigate whether Serpine1 exists as a pseudogene in chicken or whether it was not annotated correctly in databases, Serpine1 was searched in the chicken genome using reciprocal tBLASTn. The serpin was not found and the closest gene identified by reciprocal tBLASTn against human genome referred to SERPINE2, which was actually identified in both Pubmed and Ensembl databases and referenced in Table  1. Thus, to date, we cannot ascertain that this gene is absent due to errors in genome annotation or whether it actually disappeared in chickens during evolution. Chicken ov-serpins are largely represented as 10 clade B serpins could be identified  [1, 2]. These ov-serpins are clustered on a 150 kb locus chromosome 2q (Fig. 1B ) and comprise Serpinb1, Serpinb2, Serpinb5, Serpinb6, two Serpinb10 homologs (Serpinb10, Serpinb10b/MENT), Serpinb12 and Serpinb14, Serpinb14b and Ser-pinb14c namely ovalbumin and its related genes Y (OVAY) and X (OVAX).
Three orthologues to ovalbumin are referenced in duck, flycatcher and turkey in Ensembl database but information is lacking for other oviparous species. However, it is noteworthy that we found a potential orthologous of Serpinb14c in Alligator mississippiensis (A0A0Q3ZV65), sharing 56% protein sequence identity with the chicken homolog. These ovalbumin genes adapted to oviparous species, are supposed to have lost their protease inhibitor activity  [1] , and potentially acquired specific properties adapted to the development of an embryo in an egg exposed to terrestrial environments  [8, 9]. Clade A, D, F, G and H serpins are found in the second phylogenetic tree. As shown in Fig. 3 , except clade A serpins, each of these serpins has an orthologue in both G. gallus and Bos taurus. Similarly to clade B serpins, and for both species (G.
gallus; B. taurus), duplication events led to the presence of several clade A serpins after species divergence (Fig. 4 ). Concerning G. Gallus genome, this duplication gave rise to Serpina1, a3, a4, a5, a9, a10 and a12. These serpins have similar peptidase inhibitor function and are essentially expressed by the liver. The synteny analysis (http://www. genomicus. biologie. ens. fr/) reveals that clade A genes are localized in a gene cluster in chicken chromosome 5 (Fig. 3 ). The same phenomenon is observed in B. Taurus, where all clade A serpins are localized in a unique gene cluster on chromosome 21. Clade A serpins show multiple recent duplications leading to ten   [1] , based on the high sequence identity of Serpinb10 and Serpinb10b, suggesting that they are paralogs. Accession numbers are indicated in Table  1.
expasy. org). The figure was prepared with PYMOL software. Fig. 8. Schematic representation of a fertilized egg at day 16 of incubation and serpins identified in each egg structure during chicken development. For each compartment, the size of the font roughly indicates the relative quantity of one serpin to another. Bigger letter corresponds to abundant to very abundant proteins, smaller letters correspond to low abundant proteins and normal letters to proteins with intermediate abundance. Note that compartments are not comparable, as the relative quantity intimately depends on the intrinsic composition of each compartment. *Data related to the relative abundance of serpins in the chorioallantoic membrane were not available  [71]. Serpins found amniotic fluid correspond to egg white proteins that presumably transfer to amniotic fluid from day 11 onwards (the composition of egg white remains globally unchanged during the first half of incubation  [48] ).
distinct serpin genes. Protein Name [Gallus gallus] Protein accession number Gene symbol Gene ID Gene accession number serpin peptidase inhibitor, clade A (alpha-1 antiproteinase, NP 001264422. 1 Serpina1 423434 ENSGALG00000023070 antitrypsin), member 1 precursor PREDICTED: protein Z-dependent protease inhibitor XP 015143253. 1 Serpina10 423432 ENSGALG00000020391 PREDICTED: serpin A3-4-like isoform X1 XP 015143255. 1 Serpina12 107049127 ENSGALG00000028598 PREDICTED: alpha-1-antiproteinase XP 015143260. 1 Serpina3 772339 ENSGALG00000020388 serpin peptidase inhibitor, clade A (alpha-1 antiproteinase, NP 001264421. 1 Serpina4 423433 ENSGALG00000010969 antitrypsin), member 4 precursor PREDICTED: alpha-1-antitrypsin isoform X1 XP 421344. 1 Serpina5 423435 ENSGALG00000020390 PREDICTED: angiotensinogen XP 004935550.
marinum strains for 2 h at an MOI of 10. Non-ingested bacteria were extensively washed and killed by gentamycin. Macrophages were then lysed with Triton X-100 and the cell lysates were plated onto Middlebrook 7H10 agar supplemented with 10% OADC for CFU determination. The data are presented as the mean number of ingested bacteria ± SEM from six separate experiments. For statistical significance: *p ≤ 0. 03, (Mann-Whitney U test) the following datasets were compared: mutant/wild-type versus mutant/wild-type + blocking antibodies of blocking antibodies on activity of non-infected macrophages. We found that PMA-induced ROS generation by macrophages in the presence or absence of anti-TLR2 or anti-CR3 mAbs was similar (RLU total = 1091; 1008; 915, respectively). Secondly, the nitric oxide production (measured as nitrite) was also similar (control macrophages = 0.
6  Involvement of TLR2 and CR3 in intracellular M. marinum survival. Macrophages were pretreated with blocking antibodies (anti-TLR2 or anti-CR3) for 1 h and incubated with M. marinum mutants for 2 h at an MOI of 10. Non-ingested bacteria were extensively washed and killed by gentamycin. Macrophages were lysed with Triton X-100 or maintained in culture for another 2 days and then lysed, plated on Middlebrook 7H10 agar supplemented with 10% OADC and cultured for CFU determination. The data are presented as fold increase in CFU/ml (number of bacteria 2 days post-infection divided by the number of bacteria after 2-h of phagocytosis), expressed as the mean ± SEM from six separate experiments. For statistical significance: *p ≤ 0. 03, (Mann-Whitney U test) the following datasets were compared: mutant/wild-type versus mutant/wild-type + blocking antibodies structures.
Before staining with the mAbs, crystallizable fragment receptors (FcRs) were blocked using D-PBS with 10% human AB serum for 15 min at room temperature to prevent non-specific antibody binding. Afterwards, the cells were washed twice in D-PBS with 1% FBS and stained with 5 μl PE-conjugated anti-TLR2 mAb, 20 μl PE-conjugated anti-CR3 mAb or 10 μl of the appropriate isotype control for 30 min at 4 °C. Then, the cells were washed twice in D-PBS with 1% FBS, suspended in D-PBS and analyzed at once with an FACS LSR II BD flow cytometer (Becton-Dickinson, USA) that was equipped with the BD FACS Diva Software. The results are presented as median fluorescence intensity (MFI), which correlates with the surface expression of the target molecule.
Production of ROS and NO by macrophages treated with blocking antibodies ROS production was measured using a luminol-enhanced chemiluminescence method. THP-1 cells in culture medium were distributed into 96-well black plate (Nunc) at the density of 1 × 10 5 cells/well and differentiated into macrophages (see above). Then, culture medium was removed and macrophages were pretreated with anti-TLR2 or anti-CR3 blocking monoclonal antibodies in HBSS as described above or cells left untreated. Thereafter, 1 μg/ml PMA (to initiate the oxygen burst) and 1 mM luminol and 40 U horseradish peroxidase (to enhance chemiluminescence were added to macrophages. Chemiluminescence was measured during 4 h at 5-min intervals using a Fluoroscan Ascent FL instrument (Labsystems, Helsinki, Finland). Data were acquired as relative light units (RLU), and the area under the curve of chemiluminescence versus assay time (total RLU) was calculated.
Production of NO by macrophages was determined using the Griess reagent (1:1 mix of 0. 1% N-(1-naphthyl) ethylenediamine dihydrochloride and 1% sulfanilic acid in 5% phosphoric acid) which detect a nitrite, a stable metabolite of NO. Briefly, obtained macrophages (1 × 10 5 cells/well in a 96-well plate, NUNC) were treated with blocking monoclonal antibodies in culture medium for 48 h (36 °C, 5% CO 2 ). The absorbance of culture supernatants was measured on a Multiscan RC ELISA reader (Labsystems) at wavelength of 550 nm. Nitrite concentration was calculated from a standard curve using sodium nitrite as a reference. The data were presented as nitrite concentration (μM).
Conclusion Our study provides four novel spontaneous mutation rate estimates in unicellular eukaryotes. Spontaneous mutations rates were assessed over 97-99. 5% of the nuclear genomes, they are higher in intergenic than in coding sequences and 5% of mutation events affect multiple nucleotides. Combined with previous spontaneous mutation rate estimates from 20 species, our data also provides evidence that the spontaneous mutation rate increases with the deviation of the genome GC-content from its equilibrium GC-content. MBE experiments of unicellular organisms, a colony of cells is transferred to a fresh agar plate at each bottleneck to allow the separation of the cells and the random sampling of a new cell. However, this is not possible in these pico-algae as they do not grow on the surface of gelled media, in contrast to Saccharomyces cerevisiae, Dictyostelium discoideum or Chlamydomonas reinhardtii  (Wloch et al.
Acknowledgments We are grateful to  Claire Hemon ,  Elodie Desgranges  and  Christophe Salmeron  for technical assistance with the mutation accumulation experiments from 2012 to 2015 and to the  Genomics of Phytoplankton lab  for support and stimulating discussions. We acknowledge the O. mediterraneus genome consortium for access to the complete genome data and the  GenoToul Bioinformatics platform from Toulouse, France , for bioinformatics analysis support and cluster availability. This work was funded by  ANRJCJC-SVSE6-2013-0005  to GP and SSF.
The final mutation candidates were filtered to remove low mapping quality regions (MQ < 50), low coverage regions (<5 reads), and shared mutations between all MA lines. The number of callable sites per genome above these thresholds was computed to estimate the per base pair mutation rate [97-99% of the genomes are callable (supplementary table and se- quenced with Illumina technology by GATC biotech V R (Konstanz, Germany). Two different sequencing technol- ogies were used: MiSeq for O. tauri and O. mediterraneus, and HiSeq for B. prasinos and M. pusilla. Reads from ances- tral types and MA lines were aligned to the reference ge- nomes using BWA (Li and Durbin 2010) (M. pusilla: GCA_000151265. 1; O. tauri: GCF_000214015. 2); B. prasinos (ORCAE database; Sterck et al. 2012); O. mediterraneus (S Yau et al.
1. 2 Dunan (Yonaguni) Dunan (iso 639-3 yoi), also called Yonaguni,3 is a highly endangered Japonic language of the Southern Ryukyuan branch, spoken by approximately 400 speakers located on Yonaguni Island (Okinawa prefecture, Japan). In sharp contrast with its relatives, Dunan exhibits a unusually complex verb morphology for a Japonic language, mostly due to some drastic historical changes. The verb morphology of Dunan seems be the most complex one within the whole Japonic family, and a systematic description challenges in several interesting ways morpheme-based approaches. The present work is based on original data gathered by the authors during fieldwork on Yonaguni Island between 2010 and 2014. The analysis relies on a morphological database containing more than 500 verbs obtained through either elicitation or analysis of oral text recordings.
In order to test our understanding and analysis of Dunan verb morphology, we fully formalized our description and implemented it as a finite state transducer with the xfst software  (Beesley & Karttunen 2003). The transducer obtained can be used for both generation and recognition. As a generator, it generates the complete paradigm of any verb, i. e. the set of all its possible inflectional forms together with their morphosyntactic labels. When used as a morphological analyzer, the transducer can output all morphosyntactic possible analyses of an unlabeled inflected form, i. e. its morphosyntactic features and the identity and conjugation-class membership of the underlying verbal lexeme. Preliminary 2 E. g. Ashworth 1973  on Shuri Okinawan,  Lawrence (1990)  on Nakijin Okinawan, or  Arimoto (2001)  on Dunan.
For posterior risks, calculations get slightly more complex since the posterior individual hazard must now be integrated over the (changing over time) posterior joint distribution of the individual genotype and of the random effect. Basically, all computations are slightly more intensive with random effects, but most results of Section 2. 3 remain very similar. One of the important limitations of the present work is the fact that we assume that all model parameters are known. However, it should be noted that likelihood and conditional likelihood might be easy to compute through the belief propagation which means that we basically provide all the necessary means to estimate the model parameters from actual data. In that context, it is nevertheless critical to deal efficiently with ascertainment issues: the fact that the family ending up in the database are usually precisely the one with the most severe disease family history.
Acknowledgments We would first like to thank both anonymous reviewers for their constructive comments and remarks. This work received the support of both the  Institut de Recherche en Santé Publique (IRESP)  and the  Ligue National Contre le Cancer (LNCC). Alexandra Lefebvre 's internship was funded by the  Institut Curie. We finally warmly thank  Antoine de Pauw  ( Institut Curie ) for his continuous friendly support and for suggesting the hypothetical family presented here.
Protein Expression and Purification. Transmembrane domain peptides of 22 amino acid residues (VAMP2 95-116 ) were synthesized and purified as previously reported  21. Recombinant GST-VAMP2 (rat) was produced in E. coli BL21 (DE3), solubilized in Triton-X100 containing buffer and bound to glutathione beads  62. After exchanging the detergent for 0. 8% octylglucoside, VAMP was eluted from the beads by thrombin cleavage, concentrated and further purified by FPLC (Superdex75). Purified proteins were stored at -80 °C in phosphate-buffered saline containing 0. 8% octylglucoside and 10% glycerol 26 (Supplementary Fig. 1 ). Electrophysiology. The electrophysiological recordings were performed using and EPC-10 amplifiers and the Pulse software (version 8. 30 or later; Heka Elektronik, Lambrecht, Germany).
9 nm) for at least one minute. INS-1 832/13 Cells were stimulated by the main perfusion of 35 mM KCl KRBG. PC12 cells were stimulated by local perfusion with 90 mM KCl in modified KRBG (containing 50 mM NaCl) using an electrovalve. For image treatment, the brightest pixel of the fusion event was determined as the spatial and temporal origin of the event using Matlab (Mathworks, Natick, MA, USA). Automatic detection of the fusion event is based on the pattern of VAMP2-pHL fluorescence variations during exocytosis. This is typically composed by an increase in fluorescence intensity for two frames (0. 2 s) followed by a decrease over maximally seven subsequent frames (0. 7 s). Events were detected using an advanced version of a published protocol  69  and detected events were confirmed or rejected according to visual inspection.
The imaging ellipsometer was used at an incidence angle close to the Brewster angle (54. 58°). It operates on the principle of classical null ellipsometry  70. All experiments were performed at room temperature. Image patterning was determined by fractal analysis using the Fraclac plugin of ImageJ with 12 grid positions  71. Fractal dimensions given here were obtained by box counting and the values indicated (D B ) describe the slope of the relationship ln N/ln ε where N are changes in image details and ε the scale, thus indicating how a pattern's detail changes with the scale at which it is considered. To obtain structural models, a structure deposited in the PDB (code 2KOG) was used and mutations were performed using the "mutated residue" facility in the Visual Molecular Dynamics software VMD1.
8. 6. Statistical Analysis and Data Fitting. Data were expressed as the means ± SEM. Differences between two groups were assessed by a two-tailed unpaired Student's t-test or by a one-way ANOVA using XLSTAT software followed by post-hoc tests as indicated in the legends. The null hypothesis was rejected at the level of p < 0. 05. All data generated or analysed during this study are included in this published article (and its Supplementary Information files).
Thousands of fluorescent signals can be automatically detected, tracked and quantified from a time-lapse movie of the cell surface in under an hour. This has allowed the analysis of huge datasets and thus accurately depicted CME with a high power of statistical significance  (Loerke et al. , 2009). Although a few recent studies in plants have used such algorithms  (Wang et al. , 2015) , the vast majority rely on manual analysis methods using ImageJ  (Konopka et al. , 2008; Bashline et al. , 2013; Martins et al. , 2015). This mostly involves making kymographs of a time-lapse TIRF or VAEM ; the illumination beam hits the interface of two media with different refractive indexes (for example, the glass coverslip and the sample), at a sub-critical angle. The beam is refracted toward the coverslip, which produces a 'cone' shape of illumination, where the further from the point of refraction the greater the depth of penetration.
The example images are of the same Arabidopsis root tip, expressing TPLATE-GFP, but with the angle of the illumination beam altered for each circumstance. Scale bar: 500 µm. movie and manually measuring each line of fluorescence in ImageJ. This is a very time-consuming endeavor, and can take months to produce small datasets. Furthermore, this technique is not as sensitive at detecting particles as a computer algorithm, and is open to selection bias toward the brighter or persisting signals. Manual tracking is also unsuitable for proteins which are dense on the cell surface, as there is too much noise to accurately measure single traces of proteins. Taking together that TIRF is indeed possible in plants and that plant images are suitable for automated particle detection programs, we developed a reliable method for directly studying single events of CME in plants with high temporal and spatial resolution.
49 oil immersion objective (Figure  2 ). The excitation wavelengths used were 491 nm and, or, 561 nm for GFP and tagRFP, respectively, provided by a 100-mW diode laser Toptica AOTF. An emission filter Chroma ET 405/491/561/642 was used in conjunction with a Coolsnap HQ2 camera (photometrics). For single channel analyses, images were acquired at 1 s intervals, for a duration of 5 min. The time-lapse movies were subjected to only the particle detection and tracking algorithms from the cmeAnalysis package  (Aguet et al. , 2013)  using Matlab 2013b (Mathworks). Using customized homemade Matlab scripts (available upon request), the complete raw trajectories were screened so that only tracks which had both a duration greater than 5 s and that lasted less than the total length of the movie, and tracks which were not present in the first of last five frames of the movie were included in the cell surface lifetime calculation.
The data from multiple experiments were combined to produce cell surface lifetime values. Dual channel time-lapse movies were acquired sequentially at 500 ms intervals, at an interval of 1 s per channel, for a duration of 5 min. Each channel was subjected only to the particle detection and tracking stages of the cmeAnalysis package in Matlab 2013b (Mathworks), as described above. Custom homemade Matlab scripts (available upon request) were used to examine if the raw cmeAnalysis trajectories of each channel overlapped. If a 1 frame overlap was found, the different channel trajectories were paired. Paired trajectories were subjected to the departure analysis if the appearance and disappearance of the secondary channel trajectory of each channel were within 5 s of the reference channel trajectory's appearance and disappearance  (Aguet et al.
To do this, we used only the detection and tracking parts of the cmeAnalysis package  (Aguet et al. , 2013). The cmeAnalysis package was selected for use as it outperforms other detection and tracking programs available, like uTrack and Imaris  (Mettlen and Danuser, 2014). It uses a model fitting mode of detection, which allows the use of statistical test to determine if an object fits the expected point spread function of the specific fluorophore used, and if the fluorescent intensity is statistically significant compared to the local background  (Aguet et al. , 2013). cmeAnalysis detected AP2A1-GFP spots with a very high degree of accuracy (Figure  3B ). The cmeAnalysis package can complete tracks which includes gaps in them  (Aguet et al. , 2013). This is critical to determine accurate lifetimes when weak signals can fluctuate close to the limit of detection  (Jaqaman et al.