dataesr/scientific-paragraphs-categorization huggingface dataset:
org Copyright © 2020 American Society of Plant Biologists. All rights reserved. Among the 142 putative IRT1 interacting-proteins identified by co-IP combined with mass spectrometry, we present in this table proteins that are associated to intracellular trafficking or metal homeostasis according to Gene Ontogeny (GO) annotations from The Arabidopsis Information resource (TAIR) (https://www. arabidopsis. org). The maximum number of unique peptides identified for each protein from the two independent experiments (see Supplemental Dataset S1) is indicated.
The proteasome is a key player of regulated protein degradation in all kingdoms of life. Although recent atomic structures have provided snapshots on a number of conformations, data on substrate states and populations during the active degradation process in solution remain scarce. Here, we use time-resolved small-angle neutron scattering of a deuteriumlabeled GFPssrA substrate and an unlabeled archaeal PAN-20S system to obtain direct structural information on substrate states during ATP-driven unfolding and subsequent proteolysis in solution. We find that native GFPssrA structures are degraded in a biexponential process, which correlates strongly with ATP hydrolysis, the loss of fluorescence, and the buildup of small oligopeptide products. Our solution structural data support a model in which the substrate is directly translocated from PAN into the 20S proteolytic chamber, after a first, to our knowledge, successful unfolding process that represents a point of no return and thus prevents dissociation of the complex and the release of harmful, aggregation-prone products.
The disappearance of native GFP structures, observed by SANS, correlated strongly with ATP hydrolysis, which displayed the same two distinct rates. GFP molecules aggregated strongly and instantly in the presence of PAN and ATP, whereas the addition of 20S prevented aggregation and triggered the emergence of small oligopeptide products. Importantly, our data strongly support a model in which the substrate, once successfully unfolded, is directly translocated from PAN into the tightly bound 20S core particle without intermediate release of unfolded, aggregation-prone substrates. Finally, our data are compatible with the hypothesis of a proteasome-substrate complex in which an elongated, unfolded intermediate substrate spans the PAN central channel and extends into the 20S catalytic chamber, potentially serving as a ''tether'' for the two enzymatic components until proteolysis is successfully completed.
SANS data collection, reduction, and analysis SANS data sets (https://doi. ill. fr/10. 5291/ILL-DATA. 8-03-935) were recorded on the D22 diffractometer (http://www. ill. eu/instruments-support/ instruments-groups/instruments/d22) at the European Neutron Source ILL in Grenoble, France. Reaction mixtures containing 42% D 2 O (Table  S1 ) were measured at a collimator or detector configuration of 2. 0 or 2. 8 m and allowed the matching of all hydrogenated proteins (hydrogenated PAN (h-PAN) and hydrogenated 20S (h-20S)). For the reaction mixture of d-GFP and h-PAN with 100 mM ATP, an additional independent measurement was performed at a collimator or detector configuration of 8. 0 or 8. 0 m to access lower q-values for a better characterization of the aggregates. 42% D 2 O was cho-sen because it corresponds to the experimental contrast match point of h-PAN determined in a previous study  (20).
The scattering curves were recorded with an individual exposure time of 30 s. An initial dead time of $20 s was necessary to evacuate the experimental hutch and switch on the neutron beam. A neutron wavelength of 6 A ˚was used for data collection. A total of 90 individual scattering curves were recorded during the 45-min exposure times for each condition (apart from the control experiments (Fig. S2 ) in which only 45 were collected because of the limited beam time). The corresponding buffers, empty cell, empty beam, and boron/cadmium were measured in the same collimator and detector setups to perform data reduction (correction for detector efficiency, electronic noise, and sample holder scattering). Data reduction and radial integration over a two-dimensional image were performed using standard ILL software LAMP  (35) , and the one-dimensional sample and buffer curves were presented in absolute units (cm À1 ) calibrated against the direct beam.
The scattering curves of the buffers were subtracted from the respective sample curves with the software PRIMUS from the ATSAS suite  (36). The radii of gyration R G (which report on the extension of a particle) and forward scattered intensities I(0) (which report on its molecular weight), as well as the respective errors, were extracted by using the Guinier approximation (37), i. e. , a linear fit in an ln [I(q)] vs. q 2 plot, with the same program. The validity of the Guinier approximation (R G Â q max < 1. 3) was fulfilled in all cases apart from the d-GFP and h-PAN samples in which the aggregates occurred. In this latter case, the lowermost three data points were used to extract a lower estimation of R G and I(0). The program OLIGOMER  (36)  was used to fit the experimental scattering curves of GFP in the presence of PAN and 20S by using either two (natively folded GFP and decapeptide product) or three components (natively folded GFP, decapeptide product, and partially unfolded GFP intermediate).
All fits were done in the same q-range (0. 03-0. 35 A ˚À1 ). The form factor inputs (i. e. , theoretical SANS curves) of each component in 42% D 2 O were calculated by the program CRYSON  (38)  and loaded into OLIOGMER. As a representative model of native GFP, we scored an ensemble of atomic models, including a flexible C-terminus, from previous work  (39)  and selected the model that fitted the d-GFP SANS data (Fig. 1  A) best at t ¼ 0 min. As a representative decapeptide product, the first 10 residues (N-terminus) of the same model were used. The partially unfolded intermediate was created from the native GFP model by deleting the flexible C-terminus and the first, connected b-barrel strand (up to P213) using PyMOL. Subsequently, unfolded peptide residues (1-28) and the first six connected ubiquitin residues from a blocked substrate solved by cryo-EM  (18)  were fused to P213 to yield a 247-residue GFP construct (Fig.
3 , red model). The completely unfolded GFP (Fig. S10 ) was generated from native GFP by Coot (https://www2. mrc-lmb. cam. ac. uk/personal/pemsley/coot/). Complementary to OLIGOMER, we estimated the number of independent GFP-derived species in solution (in the presence of PAN and 20S) by a singular value decomposition (SVD) analysis using the program Bio-XTAS RAW  (40). Intensities were integrated over the whole q-range and for all 30-s frames during the 45-min experimental series (Fig. S13 ). The time course of I(0) intensities, I(0, t), were fitted by using a biexponential function with the Igor Pro Software (https://www. wavemetrics. com/): Ið0; tÞ ¼ I 0 þ I 1 ½1À expðÀt=t 1 Þ þ I 2 ½1À expðÀt=t 2 Þ: (1) I 0 is an initial intensity at t ¼ 0 min; I 1 and I 2 are the weight fractions of two different exponential decay processes with characteristic time rates t 1 and t 2.
The same function was also used to fit the evolution of the radii of gyration (R G ) and the fluorescence intensities (Fig. 2 ) as well as the ATP hydrolysis (Fig. S3 ). Equation  1 was also used to describe the time behavior of the natively folded GFP population from the OLIGOMER data (Fig. 3 ; Fig. S4 ). In this case, a biexponential fit did not converge, possibly because of the limited signal and noise of the original SANS data that limit the convergence of the OLIGOMER fits and/or because of the fact that the first SANS curve did not yield a 100% population of natively folded GFP. Indeed, because of the short experimental delay (20 s, see above) until neutron acquisition is started and averaged over 30 s, the reaction is already active. Therefore, a single exponential decay function (t 2 ¼ 0) was used.
Online fluorescence measurement In parallel to all TR-SANS measurements, we carried out online fluorescence measurements. To this end, a special setup with an in situ spectrofluorometer (65000 pro; Ocean Optics, Largo, FL) was implemented on the D22 beamline at the ILL as reported previously  (20). Fiber optics (Ing enerie D eveloppement Instrument Laser; Idil Fibres Optiques, Lannion, France) were connected to the SANS quartz cell containing 310 mL of the reaction volume. The excitation was ensured by a laser diode at 405 nm, and the emission wavelengths were filtered with a passband filter centered on 500 nm with a full width at half maximum of 40 nm (Thorlabs, Newton, NJ), respectively. The fluorescence emission spectra (1 s) were recorded every 30 s, and the fluorescence data were registered as NEXUS file format in the same file as the SANS data.
1 C ). Both the I(0) intensity and the fluorescence decreased rapidly over time in a biexponential process at identical fast (t 1 z 1. 5 min) and slow (t 2 z 10 min) rates (Eq. 1; Fig. 2 C ; Table  1 ). A single exponential process could not describe the decay in a satisfactory manner (Fig. S6 ). Thus, the SANS data indicate that GFP is efficiently degraded by 20S, leading to a progressive decrease  Relative changes of R G , I(0), fluorescence and populations of GFP, and ATP consumption are given. R G , I(0) and fluorescence at 509 nm (Fig. 2 ) were extracted from the SANS curves (Fig. 1 ) and from the online fluorescence measurements. The populations were determined by fitting the SANS curves with OLIGOMER (Fig. S8 ). ATP hydrolysis was measured using a standard assay (Fig. S3 ). The characteristic time constants t 1 and t 2 were fitted in all cases using Eq.
Furthermore, an SVD analysis with the program BioXTAS RAW  (40)  suggested the presence of a maximum of two significant populations of different species (Fig. S13 ). The SANS curves could be fitted well (0. 8 < c < 1. 1) by a mixture of native GFP and the representative decapeptide by using the program OLIGOMER (Fig. S8, A-C ; (36)); correspondingly, the time dependence of the populations of both species could be determined with good statistical precision (Table  S3 , left). The GFP population disappeared in a single exponential decay process (Fig. S4 ) to a final level of 13% (volume fraction) with a characteristic time of 5 min; concomitantly, oligopeptide products built up at the same rate to reach a final level of 87% (volume fraction). The rate of GFP proteolysis (5 min) lay between the slow and fast rates of the biexponential processes of ATP hydrolysis and fluorescence decay (Table  1 ).
Does a putative, elongated intermediate substrate state exist? Recently, the structures of two eukaryotic homologs of the archaeal PAN-20S complex were solved by cryo-EM in the presence of blocked substrates  (17, 18). We combined information from the substrate-engaged structure of the yeast cryo-EM complex  (18)  and from single-molecule spectroscopy  (42)  to generate a bona fide structural model of a putative, partially unfolded GFP intermediate (see Materials and Methods). This model was added to the natively folded GFP structure and the decapeptide product to fit the TR-SANS data with the program OLIGOMER. As in the case of the two-species fits, the three-species fits showed a rapid exponential decline of native GFP over the first 10 min, accompanied by an increase of the decapeptide population occurring at the same rate (Fig.
ACKNOWLEDGMENTS The authors thank  Francesca Rossano  for assistance with offline tests (Western blots and fluorescence);  Luca Signor  for mass spectrometry experiments and data reduction;  Ziad Ibrahim and Gae ¨lle Hogrel  for discussions on SANS sample experimental conditions;  Eric Girard  for the generation of a strongly unfolded GFP molecule; and  Cristina Cocho Martinez  for IT support during the TR-SANS fluorescence experiments. We acknowledge financial support from the  French Agence Nationale de la Recherche  (grant '' PROTStretch ''  ANR-15-CE11-0026-01 ). This work used the platforms of the  Grenoble Instruct-ERIC Centre  (ISBG:  UMS 3518 CNRS-CEA-UGA-EMBL ) with support from  FRISBI  ( ANR-10-INBS-05-02 ) and  GRAL  ( ANR-10-LABX-49-01 ) within the  Grenoble Partnership for Structural Biology.
Bioinformatic and evolutionary analyses The draft genome sequences of T. pallidum A10, T. pallidum A12, T. pallidum BNK07 and T. pallidum SNK05 have been deposited at European Bioinformatics Institute (EBI) and European Nucleotide Archive (ENA) respectively under the following accession numbers: OMPM01000001-OMPM01000006; OMHU01000001-OMHU01000103, OMHW01000001-OMHW01000017, OMHZ01000001-OMHZ01000216. All other Treponema genomes (CP002103, CP016062, CP016065, CP010422, CP007548, CP002375, CP002376 and CP003902) were downloaded from the National Center for Biotechnology Information (https://www. ncbi. nlm. nih. gov/). After sequencing, reads from BNK07, A10 and A12 were mapped on the reference T. pallidum Fribourg-Blanc using CLC Genomics Workbench 7 (Qiagen) with the following thresholds: 0.
Acknowledgements We are deeply grateful to  S. Calvignac-Spencer  and  F. H. Leendertz  ( Robert Koch Institute, Berlin, Germany ) for hosting and supervising hybridization capture experiments, performing part of the sequencing efforts and carefully reading of the article. We are also grateful to  C. Michelle  ( Aix Marseille Univ, MEVI ) for her technical help. Supported by the  Institut Hospitalo-Universitaire (IHU) Méditerranée Infection, France , the  National Research Agency (ANR) ,  France under the programme 'Investissements d'avenir,' reference   ANR-10-IAHU-03 ,  Région Provence Alpes Côte d'Azur ; and European funding from  FEDER-PRIMI ,  European Regional Development Fund (ERDF).
Data Analysis and Statistical Methods RNA-seq reads were aligned to hg38 using STAR aligner. Protein-coding genes with more than 10 reads in more than three samples were used for the analysis (total samples N = 86; genes N = 13,003). DESeq2 was used to find genes differentially expressed upon treatment by the menin inhibitor in each cell line compared to DMSO or hormonal treatment (for MCF-7, LNCaP and VCaP cell lines). Genes with log2 fold change (logFC) above 1 or below -1 and adjusted p-value below 0. 1 were considered significantly regulated. GSEA analysis was run on the pre-ranked list based on logFC in prostate (VCaP and LNCaP) and breast (MCF7) cell lines. Data are available at GEO under accession number GSE136272.
The classification was set and validated by WorldClim-2 at 1 km high-resolution gridded data for the 1970-2000 baseline period and 144 stations' data over 30 to 120 years, both at monthly time steps. Climatic classes coincided with a geographical distribution in the Mediterranean ranging from the most seasonal and driest class 1 in the south to the least seasonal and most humid class 5 in the north, showing the climatic continuity from one place to another and enhancing the visibility of change trends. The MED-CORDEX ALADIN and CCLM historical and projected data at 12 and 50 km resolution simulated under the RCP4. 5 and 8. 5 scenarios for the 2070-2100 period served to assess the climate change impact on this classification by superimposing the projected changes on the baseline grid-based classification.
The primary application of RCMs has been in the development of climate change scenarios, of which we mention the ALADIN RCM (Aire Limitée Adaptation dynamique Développement InterNational) developed by Météo France and the CCLM (Cosmo Climate Limited-area Model) developed by the German Weather Service (DWD), both applied for the EURO-CORDEX and MED-CORDEX projects  (Rockel et al. , 2008; Tramblay et al. , 2013). We aim in this study to discuss the results of the individual models and not to compare their performances; such a study was carried out for EURO-CORDEX with 17 RCM models for the represen-tation of the basic spatiotemporal patterns of the European climate for the period 1989-2008  (Kotlarski et al. , 2014). This paper is structured into six sections: Sect. 1 Introduction; Sect. 2 presents the Mediterranean limits and the database; Sect.
3 the classification approach based on PCA, K-means, and the decision tree with the presentation of MED-CORDEX atmosphere-RCM climate change scenarios; Sect. 4 the results of WorldClim-2 classification of gridded indices, verification of stations and comparison to catchment indices, and classification projection and impacts under MED-CORDEX scenarios; followed by a discussion in Sect. 5 before concluding with Sect. 6. 2 Study area and database.
Catchments Since the geographic extent of the study is very wide, the delimitation of catchments was imported from international references. The European Commission, using the Joint Research Centre (JRC), has done extensive and elaborate work on the delimitation of catchments in Europe and some adjacent countries as part of the Catchment Characterization and Modelling (CMM) project (de  Jager and Vogt, 2010). For catchments in the Middle East and northern Africa, catchments from HydroSHEDS, the World Wildlife Fund's project, were used  (Lehner and Grill, 2013). According to these databases, the total number of catchments extracted at their main stem outlet to the Mediterranean coastline and exceeding 1 km 2 is 3681, covering a total area of 1 781 645 km 2. It should be noted that the Nile was omitted for its extent 3500 km to the south of the Mediterranean.
Climatic data Three types of monthly climatic data were used in this study: (1) WorldClim-2 new 1 km spatial resolution climate surface data  (Fick and Hijmans, 2017) , (2) time series of 144 stations from the NOAA database of 20 different Mediterranean countries covering a period of 30 to 120 years used for validation purposes, and (3) MED-CORDEX historical and projected data simulated under the RCP4. 5 and 8. 5 scenarios for future projections  (Tramblay et al. , 2013). 1. WorldClim-2 new 1 km spatial resolution climate surface data, which consist of long-term average monthly temperature and precipitation, solar radiation, vapour pressure, and wind speed data, aggregated across a target temporal range of 1970-2000, using data from 9000 to 60 000 weather stations  (Fick and Hijmans, 2017).
The WorldClim-2 database is a refined and expanded version of the 2005 WorldClim-1 database  (Hijmans et al. , 2005). This database covers the whole study area; thus, climatic classification of Mediterranean catchments was possible. The WorldClim-2 database was built over 23 regions with different coverage for each parameter. For the precipitation an overlap of three regions covered the Mediterranean area, with a total of 10 410 stations for the three regions (western Europe n = 3730; eastern Europe n = 3632; northern Africa n = 3048). For average temperature, the Mediterranean was covered by one region (eu1) with number of stations n = 1760; n = 1627 for maximum temperature and n = 1626 for minimum temperature;    2  and Fig. 2 , reflecting the wide variability of mean annual precipitation ranging between 5 ("Jabal el Aswad desert in Libya") and 3000 mm (Kobarid in Slovenia) and mean annual temperature ranging between -14 • C (Mont Blanc, Alps, France) and +26 • C (Karak, Jordan), where some catchments receive 50 times the amount of precipitation more than others while being 4 times colder.
2. 144 ground weather station data covering the whole study area served to validate the Mediterranean climate classification, with 105 stations located within catchment boundaries and 39 outside. Also, 102 of these stations were located within Köppen's (Csa) and (Csb) Mediterranean climate, and 42 outside. These stations belong to the Global Historical Climatology Network -GHCN  (Menne et al. , 2012)  -and are recognized by the World Meteorological Organization (WMO); they are available for free access on the portal of the National Administration of Oceans and Atmosphere of the United States (NOAA). The average length of data series is 60 years and ranges between 30 and 120 years at a monthly time step. The 1960-1990 period is common to all stations. The data quality was verified (i. e. ellipse of  Bois, 1987) , and only complete hydrological years were retained for indices calculation.
Hydrology-driven climatic indices The hydrology-driven independent climatic indices were chosen subjectively and developed at the grid and catchment scales from WorldClim-2 monthly average data and divided into four groups to highlight the Mediterranean seasonality and precipitation intermittence hypothesis of the climate and its corresponding hydrological response. The climatic indices were inspired by Köppen's definition of Mediterranean climates to emphasize the precipitation and temperature variability between seasons and from the components of the water balance in its general form P = S + U + V (P : precipitation, S: surface runoff, U : underground runoff, V : evapotranspiration) to highlight the link between climate and hydrology. Hence Group I and II indices I s , P 25 % , P 75 % , and I Decal characterize Mediterranean precipitation P in its seasonality and monthly distribution.
Principle component analysis Principal component analysis is widely applied to reduce the dimensionality of datasets and keep the most representative and uncorrelated variables. This section presents a brief description of the method along with some of their applications in hydrology. For an extensive mathematical description and demonstration of these methods, we advise the reader to consult  Krzanowski (1988)  and Jollife  (2002). PCA was first introduced by Karl Pearson  (Pearson, 1901 ) and then developed by Harold Hotelling  (Hotelling, 1933 ). Hotelling's (1933)  motivation is that there may be a smaller fundamental set of independent variables which determine the values and conserve the maximum amount of information of the original variables  (Jolliffe, 2002). This is achieved by transforming a vector of p random variables to a new set of variables, named principal components (PCs), by looking for a linear function of the elements with maximum variance and next looking for another linear function uncorrelated with the first and having maximum variance and so on up to p PCs.
Decision tree A decision tree is a set of distance criteria or questions in the form of a hierarchy that leads to an intended classification  (Breiman, 1984). To classify new points or stations or to reproduce the classification on another dataset, it suffices to define the distance criterion to the various kernels of the climatic classes by predicting values of a dependent variable based on values of predictor variables from a reference classification. This procedure provides validation tools for exploratory and confirmatory classification analysis. We generated a decision tree based on the distances to the clusters' kernels obtained from the gridded indices classification. The aim of this decision tree is to easily reproduce the classification with the same kernels rather than to repeat the whole classification process, which will modify the clusters and their kernels.
Adopted methodology The proposed methodology consisted first in calculating the grid-based climatic indices using WorldClim-2 monthly data, second in reducing the number of indices with the PCA, and third in classifying it using K-means clustering. The gridded indices classification was later verified on the ground station indices and then compared to the catchment-scale-averaged data for future hydrological applications. In addition, a hierarchical decision tree was constructed to avoid repeating the whole process when classifying projected indices. All PCAs, K-means, and decision trees were calculated using SPSS software. Projected indices under RCP scenarios were calculated and classification evolutions were then deduced.